Rapid change in sample temperature for measurement

Abstract

In addition, the present invention provides an apparatus and method capable of rapidly changing or cycling (i.e., heating and cooling) the temperature of a sample with high speed, less heating energy, high energy efficiency, a compact and simplified device (e.g., a handheld device), easy and rapid operation, and / or low cost.

Description

[0001] Cross - reference

[0002] This application claims the benefit of U.S. Provisional Patent Application No. 62 / 510,063, filed May 23, 2017; International Application No. PCT / US2018 / 017307, filed Feb. 7, 2018; International Application No. PCT / US2018 / 018108, filed Feb. 14, 2018; International Application No. PCT / US2018 / 018405, filed Feb. 15, 2018; and International Application No. PCT / US2018 / 028784, filed Apr. 23, 2018, each of which is hereby incorporated by reference in its entirety for all purposes. Field of the Invention

[0003] In addition, the present invention relates to devices and methods for performing biological and chemical assays, and more particularly to devices and methods having rapid sample temperature changes, rapid assays, and ease of use. Background of the Invention

[0004] In certain chemical, biological, or medical assays, rapid sample temperature changes or rapid thermal cycling are required (e.g., for polymerase chain reaction (PCR) or isothermal amplification of nucleic acids).

[0005] To increase the sample thermal cycling speed (i.e., shorter thermal cycling times), small sample sizes, small thermal cycling chambers, and rapid heating elements with small thermal masses (e.g., radiant heaters or electrical heaters) have been described in the prior art. However, there is still a need for devices and methods that can improve the speed, energy efficiency, device size and weight, operating procedures and time, power consumption, and / or cost of rapid sample thermal cycling. Such improvements can have significant economic benefits.

[0006] An object of the present invention is to meet these needs. The present invention also provides useful devices and methods for isothermal nucleic acid amplification. Summary of the Invention

[0007] The following brief summary is not intended to include all features and aspects of the present invention.

[0008] In addition, the present invention provides devices and methods capable of rapidly changing or cycling (i.e., heating and cooling) the sample temperature at high speed, with less heating energy, high energy efficiency, a compact and simplified device (e.g., a handheld device), easy and rapid operation, and / or low cost.

[0009] The present invention has experimentally achieved cycling of the sample temperature between 95°C and 55°C within one second or less.

[0010] The present invention has six novel aspects: (1) devices and methods that allow for rapid thermal cycling, (2) devices and methods that allow for uniform sample thickness and mechanical stability of the sample holder for processing, (3) simple operation, (3) devices and methods for performing real-time PCR, (4) biochemistry, and (5) a smartphone-based system.

[0011] To rapidly thermally cycle the temperature of a sample or a portion thereof, the thermal mass and lateral heat must be reduced.

[0012] Radiative heating and cooling are preferred. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Those skilled in the art will understand that the drawings described below are for illustrative purposes only. The drawings are not intended to limit the scope of the present invention in any way. In some cases, the drawings are not drawn to scale. In the figures presenting experimental data points, the lines connecting the data points are for guiding the observation of the data only and have no other meaning.

[0014] FIG. 1 shows a schematic diagram of certain components of a system for changing the temperature of a sample and for monitoring signals from the sample according to some embodiments.

[0015] FIG. 2A shows an embodiment of a device having a heating layer separated from a cooling layer according to some embodiments.

[0016] FIG. 2B shows an embodiment of a heating layer having a contact cooling layer according to some embodiments.

[0017] FIGS. 3A and 3B respectively show a perspective view and a cross-sectional view of a combined heating / cooling layer on the outer surface of a plate according to some embodiments.

[0018] FIG. 4A shows a perspective view and a cross-sectional view of a device in an open configuration according to some embodiments.

[0019] FIG. 4B shows a perspective view and a cross-sectional view of the device when the sample holder is in a closed configuration according to some embodiments.

[0020] FIG. 5 shows a top view of a device according to some embodiments.

[0021] FIG. 6A shows a perspective view of a system when the device (the sample holder of the system) is in an open configuration according to some embodiments.

[0022] FIG. 6B shows a cross-sectional view of the system when the sample holder is in a closed configuration according to some embodiments.

[0023] FIG. 7 shows a cross-sectional view of a system according to some embodiments showing additional elements that facilitate temperature change and control.

[0024] Figures 8A and 8B respectively show a perspective view and a cross-sectional view of a device having multiple sample contact areas according to some embodiments.

[0025] Figure 9 shows a cross-sectional view of a device demonstrating how to add and squeeze a sample according to some embodiments.

[0026] Figure 10 shows a cross-sectional view of a device demonstrating the PCR process according to some embodiments.

[0027] Figures 11A and 11B respectively show a top view and a cross-sectional view of a heating layer on a plate of a device according to some embodiments.

[0028] Figures 12A and 12B show cross-sectional views of a device having a first plate, a second plate, and a heating / cooling layer according to some embodiments.

[0029] Figure 13 shows a cross-sectional view of a system for rapidly changing the temperature of a sample including a heating source using fibers according to some embodiments.

[0030] Figure 14 shows a cross-sectional view of a system for rapidly changing the temperature of a sample including a heating source using a lens according to some embodiments.

[0031] Figures 15A and 15B respectively show a top view and a side view of a device having separate heating elements according to some embodiments.

[0032] Figures 16A and 16B respectively show a perspective view and a side view of an optical waveguide for guiding electromagnetic waves (such as light) from a heating source according to some embodiments.

[0033] Figure 17 shows a perspective view of an optical waveguide according to some embodiments.

[0034] Figures 18A and 18B respectively show a side view and a top view of a sample device heated by a heat source according to some embodiments.

[0035] Figure 19 shows a schematic side view of a device having a lens for focusing light from a heat source according to some embodiments.

[0036] Figure 20 shows experimental absorption spectra of different materials according to some embodiments.

[0037] Figure 21 shows experimental thermal cycling data according to some embodiments.

[0038] Figure 22 shows experimental data on the effect of the area of the heating / cooling layer on heating and cooling times according to some embodiments.

[0039] Figure 23 shows experimental data on heating and cooling times relative to the area dimensions of the heating / cooling layer according to some embodiments.

[0040] FIG. 24A shows experimental data of the relationship between heating time and the thickness of the heating / cooling layer according to some embodiments.

[0041] FIG. 24B shows experimental data of the relationship between cooling time and the thickness of the heating / cooling layer according to some embodiments.

[0042] FIG. 25A shows experimental data of the relationship between heating time and the distance between the heating / cooling layer and the sample according to some embodiments.

[0043] FIG. 25B shows experimental data of the relationship between cooling time and the distance between the heating / cooling layer and the sample according to some embodiments.

[0044] FIG. 26A shows experimental data of the relationship between heating time and the thickness of the sample layer according to some embodiments.

[0045] FIG. 26B shows experimental data of the relationship between cooling time and the thickness of the sample layer according to some embodiments.

[0046] FIG. 27A shows experimental data of the relationship between heating time and the power of the heating source according to some embodiments.

[0047] FIG. 27B shows experimental data of the relationship between cooling time and the power of the heating source on the sample according to some embodiments.

[0048] FIG. 28A shows experimental data of the relationship between heating time and different heating / cooling layer materials according to some embodiments.

[0049] FIG. 28B shows experimental data of the relationship between cooling time and different heating / cooling layer materials according to some embodiments.

[0050] FIG. 29A shows a schematic diagram of a device with spherical spacers according to some embodiments.

[0051] FIG. 29B shows a schematic diagram of a device with columnar spacers according to some embodiments.

[0052] FIGS. 30A and 30B respectively show a top view and a side view of a device on a support according to some embodiments.

[0053] FIG. 31 shows experimental data of the effect of placing a device on a device support and / or a device adapter on heating and cooling times according to some embodiments.

[0054] DETAILED DESCRIPTION OF EXEMPLARY EMBODIMENTS

[0055] The following detailed description shows some embodiments of the present invention by way of example and not limitation. The chapter headings and any subheadings used herein, if any, are for organizational purposes only and should not be construed as limiting the subject matter described in any way. The table of contents under the chapter headings and / or subheadings is not limited to the chapter headings and / or subheadings, but applies to the entire description of the present invention.

[0056] Any reference to a publication is for disclosure prior to the filing date and should not be construed as an admission that the claimed invention is not entitled to antedate such publications by virtue of prior invention. In addition, the provided publication dates may be different from the actual publication dates, which may require separate confirmation.

[0057] It should be noted that the drawings are not intended to show the elements in strict proportion. For clarity, some elements are shown enlarged in the figures. The dimensions of the elements should be depicted in accordance with the description provided herein and incorporated by reference.

[0058] Definitions

[0059] The term "sample thermal cycler" or "thermal cycler" refers to a device that can raise and cool the temperature of a sample, and if desired, the device can repeatedly heat and cool the sample between two temperatures.

[0060] The term "sample thermal cycling" or "thermal cycling" refers to repeatedly raising and cooling the temperature of a sample.

[0061] The term "sample thermal cycling" or "thermal cycling" refers to a cycle in which the sample temperature is raised to a higher temperature and then cooled back to the original temperature.

[0062] The term "sample thermal cycle time" or "thermal cycle time" refers to the time required to perform a given number of thermal cycles.

[0063] The term "sample thermal cycle rate" or "thermal cycle rate" refers to the rate at which thermal cycling is performed.

[0064] The "thermal mass" of a material refers to the energy required to heat the material by one degree when there is no other energy loss. Thus, the thermal mass of a material is equal to the specific heat per unit volume multiplied by the volume of the material.

[0065] The term "ratio of thermal conductivity to capacity" refers to the ratio of the thermal conductivity of a material to its heat capacity. For example, at about room temperature, the ratio of thermal conductivity to capacity of gold is 1.25 cm 2 / sec (square centimeters per second) and the ratio of thermal conductivity to capacity of water is 1.4×10 -3 cm 2 / sec.

[0066] The term "wasted energy" refers to the energy supplied to a sample holder that is not used for directly heating the relevant sample.

[0067] As used herein, the terms "a" and "an" are to be understood as meaning "at least one" unless otherwise expressly stated.

[0068] As used herein, the term "about" generally refers to a range that is greater than or less than the stated value by 15% in the context of a particular use. For example, "about 10" shall include the range from 8.5 to 11.5.

[0069] The term "sample holder support" refers to the device to which the sample holder is physically attached and mechanically supported by the device.

[0070] As used herein, the term "disposable" generally refers to a device that is designed to be discarded after limited use (e.g., in terms of the number of reactions, thermal cycles, or time) rather than being used repeatedly indefinitely.

[0071] The term "nucleic acid amplification" refers to the production of one or more replicated copies of an existing nucleic acid.

[0072] The term "nucleic acid amplification cycle" refers to the entire set of steps used to perform a single round of nucleic acid amplification.

[0073] The term "template" refers to the nucleic acid to be amplified.

[0074] The term "amplification product" refers to the replicated copy of an existing nucleic acid produced from the template during nucleic acid amplification.

[0075] The term "black paint" refers to paint that appears black to the human eye under sunlight irradiation.

[0076] The terms "cooling gas" or "cooling liquid" refer respectively to the gas phase or liquid phase used to remove thermal energy, for example, from a sample, from a sample holder, from a material, or from a region.

[0077] As used herein, the term "mechanical contact" generally refers to the contact formed between one or more materials, where the materials are in physical contact.

[0078] The term "thermal path" refers to the distance through which thermal energy is transferred from one location to another.

[0079] The terms "relevant sample" or "relevant sample volume" refer to the volume of the sample that is heated and / or cooled to the desired temperature during thermal cycling, and the relevant sample can be a part or the entire volume of the sample on the sample holder, and there is no fluid separation between a part of the sample and the rest of the sample.

[0080] The term "high-K material" refers to a material with a thermal conductivity (K) equal to or greater than 50 W / (m·K) (e.g., gold: -314 W / (m·K) and graphite -80 W / (m·K) are high-K materials).

[0081] The term "low-K material" refers to a material with a thermal conductivity (K) equal to or less than 1 W / (m·K) (e.g., water (-0.6 W / (m·K)) and plastic (-0.2 W / (m·K)) are low-K materials).

[0082] The terms "cooling time in the thermal cycle" and "cooling cycle time" are interchangeable.

[0083] The terms "heating time in the thermal cycle" and "heating cycle time" are interchangeable.

[0084] The term "heating zone" refers to (a) the heating layer when the heating layer is a layer separated from the cooling layer; or (b) the heating area when heating and cooling use the same layer; the heating zone is directly heated by a heating source.

[0085] The term "direct heating" refers to the energy input to the area. For example, for a heating zone heated by an LED heating source, the LED heating source projects light above the heating zone. For a heating zone heated by an electric heating source, the electric heating source sends current to the heating zone to generate heat in the heating zone.

[0086] The term "cooling zone" refers to (a) the cooling layer when the cooling layer is a layer separated from the heating layer; or (b) the cooling area when cooling and heating use the same layer. Unless otherwise specified, the cooling zone contains a material with a thermal conductivity of 50 W / m- or greater.

[0087] The term "the heating layer is heated by a heating source" means "the heating layer or the heating zone of the heating / cooling layer is heated by a heating source".

[0088] The term "non-sample material" refers to the material on the sample holder outside the relevant sample volume.

[0089] The term "wasted heating energy" refers to the energy that must be provided to the non-sample material and non-relevant samples to heat the relevant sample volume to the desired temperature.

[0090] The "average linear dimension" of a region is defined as the length equal to the area multiplied by 4 and then divided by the perimeter of the area. For example, if the region is a rectangle with width w and length L, the average value of the linear dimension of the rectangle is 4*W*L / (2*(L + W)) (where "*" represents multiplication and " / " represents division). According to this definition, for a square with width W, the average linear dimension is W, and for a circle with diameter d, the average linear dimension is d.

[0091] The term "lateral" refers to a direction parallel to the plate of the sample holder.

[0092] The term "vertical" refers to a direction perpendicular to the plate of the sample holder.

[0093] The "period" of an array of periodic structures refers to the distance from the center of a structure to the center of the nearest adjacent identical structure.

[0094] The terms "smartphone" or "mobile phone", which are used interchangeably, refer to a type of phone having a camera and communication hardware and software that can use the camera to take images, manipulate the images taken by the camera, and transmit data to a remote location. In some embodiments, the smartphone has a flash.

[0095] Unless otherwise specified, the term "heating layer" or "heating zone" refers to a layer of material that includes at least one layer of material having a thermal conductivity of 50 W / m-K or greater.

[0096] The term "heating volume" refers to the volume of the material to be heated. "Heated sample volume" refers to the volume of a portion of the heated sample.

[0097] The term "cooling layer" refers to a thermally radiative cooling layer having a high thermal conductivity and a large surface thermal radiation capacity, where the large surface thermal radiation capacity is at least 50% of the surface thermal radiation capacity of a black body.

[0098] The terms "lateral dimension" or "lateral area" of a sample inside a sample holder for heating and cooling refer to the lateral dimension or lateral area of the portion of the sample that is heated to a desired temperature.

[0099] The term "plate" refers to a self-standing plate, except when two plates are in a "closed configuration", in which case the two plates are closed together and separated by spacers (in which case the pair of plates is self-standing). The term "self-standing" means that the central region of the plate has no support. For example, when two plates are in a closed configuration and a sample is between the plates. The central region of the pair of plates has no mechanical support, and only air contacts the outer surfaces of the plates.

[0100] Principle of operation

[0101] One aspect of the present invention is to reduce the thermal cycle time, reduce the heating energy used for such cycles, increase the energy efficiency, and reduce the total power consumption.

[0102] The thermal cycle time (speed), heating energy, energy efficiency, and power consumption are related. When more heating energy is required to raise the temperature of a given sample, more energy must be removed when cooling the sample, which in turn requires more time and / or more energy for cooling.

[0103] Many thermal cyclers in the prior art require a large amount of heating energy to maintain the sample holder (e.g., plastic chamber walls) rather than the sample; use lateral heat conduction through the large thermal mass and poor heat conduction materials of the sample holder as the main cooling channel for cooling the sample (note that the material needs to absorb and release energy for heat conduction); use conduction cooling as the main cooling method, and / or use additional cooling gas or moving cooling blocks. These methods result in problems such as long thermal cycling time, high thermal energy, low energy efficiency, bulky equipment, and / or high cost.

[0104] Based on theoretical and experimental studies, the present invention provides solutions to certain drawbacks in sample thermal cycling in the prior art.

[0105] To demonstrate the working principle of the present invention, let's look at the energy components in heating and cooling the sample. There are three common energy components for heating and cooling: (i) one involving thermal mass (i.e., the ability of a material to absorb and store energy); the larger the thermal mass, the more energy needs to be added for heating and the more energy needs to be removed during cooling, (ii) heat loss caused by thermal radiation, and (iii) heat loss caused by heat conduction / convection. For rapid heating, all three energy components need to be small. However, for rapid cooling, the first energy component needs to be small, but at least one of the last two energy components needs to be large.

[0106] Through theoretical and experimental studies, the present invention is based on certain designs that can balance and / or optimize the three energy components for rapid heating and cooling. In particular, in certain embodiments, the present invention reduces the thermal mass that must be heated during thermal cycling, limits lateral heat conduction, and uses radiative heat loss as the main way to remove energy from the heated sample.

[0107] According to the present invention, the cooling of the sample is significantly by radiative cooling rather than by conduction cooling. Thus, during thermal cycling, most or most of the non-sample materials on the sample holder do not absorb and release as much energy as in a heat conduction-dominated system.

[0108] One aspect of the present invention provides devices and methods for reducing the heating of non-sample materials on the sample holder.

[0109] Another aspect of the present invention provides devices and methods for reducing lateral heat conduction through the large thermal mass and poor heat conduction materials on the sample holder.

[0110] Another aspect of the present invention provides devices and methods for using radiative cooling as the main cooling channel to cool the sample.

[0111] Another aspect of the present invention provides an apparatus and method for placing spacers between plates (i.e., walls) sandwiching a sample. The spacers provide good sample uniformity over a large area, even when the plates are thin (e.g., 25 μm thick) and flexible. In the absence of spacers, it is difficult to obtain a uniform sample thickness when the two plates defining the sample become very thin.

[0112] Another aspect of the present invention provides an apparatus and method for making the operation of the apparatus simpler.

[0113] According to the present invention, radiative cooling uses a layer of material (in terms of material and shape) that has good radiative cooling characteristics during cooling and low thermal mass (and thus low heating energy) during heating.

[0114] According to the present invention, the sample holder is configured to limit / minimize conductive cooling.

[0115] According to the present invention, the sample thickness of the sample chamber wall thickness, the first and second plates (facing each other) are configured to reduce lateral heat conduction (i.e., in the direction of the plates).

[0116] According to the present invention, in some embodiments, the radiative cooling layer is the same heating / cooling layer as the heating layer, but the ratio of the cooling zone to the heating zone, the material properties, and the material thickness and geometry are configured such that the heating / cooling layer has low thermal mass during heating and a high radiative cooling rate.

[0117] Another object of the present invention is to perform one cycle of sample temperature change (e.g., from 95 °C to 55 °C) in a few seconds or even a few seconds (e.g., 0.7 seconds).

[0118] Another aspect of the present invention is to provide useful apparatus and methods for isothermal nucleic acid amplification, where the sample temperature needs to be raised from ambient temperature to an elevated temperature (i.e., 65 °C) and held for a period of time (i.e., 5 - 10 minutes). One aspect of the present invention is rapid heating, using less energy, and making the apparatus compact, lightweight, and portable.

[0119] The thermal mass of the card and the sample is minimized to reduce the energy required for heating and the energy required for cooling.

[0120] Another aspect of the present invention is that in certain embodiments, only a small portion of the sample is heated and / or cooled.

[0121] Another aspect of the present invention is that it uses a thin layer of high thermal conductivity, the area size of which is larger than the area size of the relevant sample region.

[0122] Another aspect of the present invention is to use a thin high - thermal - conductivity layer with an area size larger than the area of the heating zone.

[0123] Another aspect of the present invention provides an apparatus and method for reducing heating of non-sample materials on a sample holder.

[0124] Another aspect of the present invention provides an apparatus and method for reducing lateral heat conduction in a large thermal mass and poor thermal conductivity material on a sample holder.

[0125] Another aspect of the present invention provides an apparatus and method for cooling a sample using thermal radiation cooling as a primary cooling channel.

[0126] Another aspect of the present invention is that it can achieve rapid thermal cycling without using a cooling gas.

[0127] The thermal mass of the card and the sample is minimized to reduce the energy required for heating and the energy required for cooling.

[0128] Another aspect of the present invention is to regulate radiative cooling and convective cooling for rapid cooling.

[0129] A heat sink for radiative cooling and / or convective cooling is used for rapid cooling.

[0130] An embodiment of the sample thermal cycling device of the present invention (as shown in FIG. 1) includes (i) a sample holder, which is referred to as an "RHC (Rapid Heating and Cooling) card" or a "sample card", which allows rapid heating and cooling of a sample on the card; (ii) a heating source; (iii) an additional heat sink (optional); (iv) a temperature control system, and (v) a signal monitoring system (optional). The temperature control system and the signal monitoring system are not explicitly shown in FIG. 1, but can be used to control the output of the heating source. In some embodiments, a signal sensor is included to detect an optical signal from the sample on the sample holder. Note that some embodiments of the present invention may only have one or several components shown in FIG. 1.

[0131] FIGS. 2A and 2B show cross-sectional views of two embodiments of the apparatus of the present invention. FIG. 2A shows an embodiment including a separate heating layer (112-1) and a separate cooling layer (112-2), where the heating layer (112-1) is on the outer surface of one plate, and the cooling layer (112-2) is on the outer surface of another plate. FIG. 2B shows an embodiment including a heating layer (112-1) and a cooling layer (112-2), where the heating layer (112-1) and the cooling layer (112-2) are structurally different but in contact with each other, and both layers are on the outer surface of one of the plates.

[0132] SH-1 A detailed description of an embodiment of the RHC card of the present invention is an apparatus for rapidly changing the temperature of a fluid sample, comprising:

[0133] A first plate (10), a second plate (20), a heating layer (112-1) and a cooling layer (112-2), wherein:

[0134] Each of the first plate and the second plate has a sample contact area on its respective inner surface for contacting a fluid sample; wherein the sample contact areas face each other, are separated by an average spacing of 200 μm or less, and are capable of sandwiching the sample therebetween;

[0135] Heating layer:

[0136] Is positioned on the inner surface, outer surface or inside of one of the plates,

[0137] Is configured to heat a relevant volume of the sample, wherein the relevant volume of the sample is part or all of the sample being heated to a desired temperature; and

[0138] Cooling layer:

[0139] Is located on the inner surface, outer surface or inside of one of the plates;

[0140] Is configured to cool a relevant sample volume; and

[0141] Contains a material layer with a thermal conductivity to heat capacity ratio of 0.6 cm 2 / sec or greater;

[0142] Wherein the distance between the cooling layer and the surface of the relevant sample volume is zero or less than the distance configured such that the thermal conductivity per unit area between the cooling layer and the surface of the relevant sample volume is equal to 70 W / (m 2 ·K) or greater; and

[0143] Wherein, in some embodiments, the heating layer and the cooling layer are the same material layer with a heating zone and a cooling zone, and wherein the heating zone and the cooling zone can have the same area or different areas.

[0144] Another detailed description of an embodiment of the RHC card (sample holder) of the present invention, SH-2, is a device for rapidly changing the temperature of a fluid sample, comprising:

[0145] A first plate (10), a second plate (20), a heating layer (112-1) and a cooling layer (112-2), wherein:

[0146] Each of the first plate and the second plate has a sample contact area on its respective inner surface for contacting a fluid sample; wherein the sample contact areas face each other, are separated by an average separation distance of 200 μm or less, and are capable of sandwiching the sample therebetween;

[0147] Heating layer:

[0148] is located on the inner surface, outer surface or inside of one of the plates,

[0149] is configured to heat a relevant volume of the sample, wherein the relevant volume of the sample is part or all of the sample being heated to a desired temperature; and

[0150] Cooling layer:

[0151] is located on the inner surface, outer surface or inside of one of the plates;

[0152] is configured to cool the relevant sample volume; and

[0153] includes a material layer having a thermal conductivity to heat capacity ratio of 0.6 cm 2 / sec or greater, wherein the layer with the high thermal conductivity to heat capacity ratio has an area greater than the lateral area of the sample volume;

[0154] wherein the distance between the cooling layer and the surface of the relevant sample volume is zero or less than the distance configured such that the thermal conductivity per unit area between the cooling layer and the surface of the relevant sample volume is equal to 150 W / (m 2 ·K) or greater; and

[0155] wherein, in some embodiments, the heating layer and the cooling layer are the same material layer having a heating zone and a cooling zone, and wherein the heating zone and the cooling zone can have the same area or different areas.

[0156] As shown in FIGS. 3A and 3B, in some embodiments of the present invention, the heating layer and the cooling layer are combined into one layer (heating / cooling layer), creating a heating zone and a cooling zone, wherein the cooling zone is larger than the heating zone. The sample card 100 (also referred to as the "RHC card") can include two thin plates (10, 20) that sandwich a fluid sample (90) therebetween, and the heating / cooling layer (112) is located below the sample, and the heating / cooling layer (112) is heated by a heating source located away from the card. According to one embodiment, at the edge of the sample, there is no wall for containing the sample, but the edge of the sample does not flow due to the capillary force that maintains the shape of the edge of the fluid sample.

[0157] As shown in FIG. 4A, according to one embodiment, the plates 10 and 20 can have inner surfaces 11 and 21 separated by a spacer 102. When the device is ready to receive the sample (e.g., in the open position), the spacer 102 can be large. FIG. 4B shows the closed configuration of the device 100, where the spacer 102 is made smaller (e.g., less than about 200 μm) to sandwich the sample 90 between the plates 10 and 20. In this embodiment, the heating / cooling layer 112 is located on the outer surface 22 of the plate 20.

[0158] Another detailed description of an embodiment of the RHC card of the present invention is a device for rapidly changing the temperature of a fluid sample, comprising:

[0159] A first plate (10), a second plate (20), and a heating / cooling layer (112), wherein:

[0160] The first plate (10) and the second plate (20) face each other and are spaced apart from each other by a certain distance;

[0161] Each plate has a sample contact area on its respective inner surface (11, 21) for contacting the fluid sample; wherein these sample contact areas face each other, contact the sample, define the sample therebetween, and have an average spacing (102) from each other, and the sample;

[0162] The heating / cooling layer (112) is located on the outer surface (22) of the second plate (20); and

[0163] The heating / cooling layer is configured to include a heating zone and a cooling zone; wherein the heating zone is configured to heat the fluid sample, and the cooling zone is configured to cool the sample by radiative cooling;

[0164] Wherein the heating zone is configured to receive heating energy from a heating source and has an area smaller than the total area of the heating / cooling layer; and

[0165] Wherein at least a part of the heating zone of the heating layer overlaps with the sample area.

[0166] SH-4. A device for rapidly changing the temperature of a fluid sample, comprising:

[0167] A first plate (10), a second plate (20), a heating layer (112-1), and a cooling layer (112-2), wherein:

[0168] The first plate and the second plate are movable relative to each other into different configurations;

[0169] Each first plate and second plate has a sample contact area on its respective inner surface for contacting the fluid sample; wherein the sample contact areas face each other, are separated by an average spacing of 200 μm or less, and are capable of clamping the sample therebetween;

[0170] The heating layer:

[0171] Is positioned on the inner surface, outer surface, or inside of one of the plates,

[0172] Is configured to heat the relevant volume of the sample, wherein the relevant volume of the sample is part or all of the sample being heated to a desired temperature; and

[0173] The cooling layer:

[0174] is located on the inner surface, outer surface or inside of one of the plates;

[0175] is configured to cool a related sample volume; and

[0176] includes a material layer having a ratio of thermal conductivity to heat capacity of 0.6 cm 2 / sec or greater;

[0177] wherein, one of the configurations is an open configuration, wherein: the two plates are partially or completely separated and the average spacing between the plates is at least 300 μm;

[0178] wherein, another of the configurations is a closed configuration, which is configured after depositing a fluid sample on one or both of the sample contact areas in the open configuration; and in the closed configuration: at least a portion of the sample is defined as a layer by the two plates, wherein the average sample thickness is 200 μm or less.

[0179] SH-5. A device for rapidly changing the temperature of a fluid sample, comprising:

[0180] a first plate (10), a second plate (20), a spacer, a heating layer (112-1) and a cooling layer (112-2), wherein:

[0181] the first plate and the second plate are movable relative to each other into different configurations;

[0182] each of the first plate and the second plate has a sample contact area on its respective inner surface for contacting a fluid sample; wherein the sample contact areas face each other, are separated by an average spacing of 200 μm or less, and are capable of sandwiching the sample therebetween;

[0183] one or both of the plates include a spacer, and the spacer is fixed to the inner surface of the corresponding plate;

[0184] the spacer has a predetermined substantially uniform height equal to or less than 200 microns and a predetermined spacer spacing;

[0185] the heating layer:

[0186] is positioned on the inner surface, outer surface or inside of one of the plates,

[0187] is configured to heat a related volume of the sample, wherein the related volume of the sample is part or all of the sample being heated to a desired temperature; and

[0188] the cooling layer:

[0189] is located on the inner surface, outer surface or inside of one of the plates;

[0190] configured to cool a related sample volume; and

[0191] comprising a material layer having a thermal conductivity to heat capacity ratio of 0.6 cm 2 / sec or greater;

[0192] wherein, one of the configurations is an open configuration in which: the two plates are partially or fully separated, the spacing between the plates is not adjusted by spacers, and the sample is deposited on one or both plates; and

[0193] wherein, another of these configurations is the closed configuration configured after depositing the sample in the open configuration; and in the closed configuration: at least a portion of the sample is squeezed by the two plates into a layer of very uniform thickness, wherein the uniform thickness of the layer is defined by the sample contact surfaces of the plates and is adjusted by the plates and spacers.

[0194] In some embodiments, the heating / cooling layer (112) may be on or within the inner surface (21) of the second plate (20), rather than on the outer surface (22) of the second plate (20).

[0195] In some embodiments of all embodiments of the device, the RHC card further includes spacers positioned between the first and second plates to adjust the distance between the two plates (i.e., the plate spacing), and thus adjust the sample thickness. The spacers may allow the sample thickness between the two plates to be uniform over a large area, even when the plates are thin and flexible.

[0196] In some embodiments, there is more than one heating / cooling layer.

[0197] A. Smaller related sample volume (RE ratio)

[0198] Reducing the sample volume that should be heated or cooled to the desired temperature can shorten the heating time, cooling time, and heating power in the thermal cycle. The reduction of the sample volume in the thermal cycle can be achieved by (a) reducing the entire sample volume or (b) heating only a portion of the sample on the sample holder. The term "related sample" or "related sample volume" refers to the volume of the sample that is heated and / or cooled to the desired temperature during the thermal cycle, and the related sample can be a portion or the entire volume of the sample on the sample holder, and there is no fluid separation between a portion of the sample and the rest of the sample.

[0199] In some embodiments, the relevant volume of the sample is 0.001 μl, 0.005 μl, 0.01 μl, 0.02 μl, 0.05 μl, 0.1 μl, 0.2 μl, 0.5 μl, 1 μl, 2 μl, 5 μl, 10 μl, 20 μl, 30 μl, 50 μl, 100 μl, 200 μl, 500 μl, 1 ml, 2 ml, 5 ml, or within a range between any two values.

[0200] In some preferred embodiments, the relevant sample volume is from 0.001 μL to 0.1 μL, from 0.1 μL to 2 μL, from 2 μL to 10 μL, from 10 μL to 30 μL, from 30 μL to 100 μL, from 100 μL to 200 μL, or from 200 μL to 1 mL.

[0201] In some preferred embodiments, the relevant sample volume is from 0.001 μL to 0.1 μL, from 0.1 μL to 1 μL, from 0.1 μL to 5 μL, or from 0.1 μL to 10 μL.

[0202] In certain embodiments, the ratio of the relevant sample to the entire sample volume (RE ratio) is 0.01%, 0.05%, 0.1%, 0.5%, 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or within a range between any two values.

[0203] In some preferred embodiments, the RE ratio is from 0.01% to 0.1%, from 0.1% to 1%, from 1% to 10%, from 10% to 30%, from 30% to 60%, from 60% to 90%, or from 90% to 100%.

[0204] To heat only a portion of the sample, in some embodiments, the area of the heating zone is only a portion of the lateral area of the sample, and this portion (i.e., the ratio of the heating zone to the lateral area of the sample) is 0.01%, 0.05%, 0.1%, 0.5%, 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99%, or within a range between any two values.

[0205] In some preferred embodiments, the ratio of the area of the heating zone to the lateral area of the sample is within a range between 0.01% and 0.1%, between 0.1% and 1%, between 1% and 10%, between 10% and 30%, between 30% and 60%, between 60% and 90%, or between 90% and 99%.

[0206] B. Local heating, with the height perpendicular to the lateral heat transfer

[0207] When a high-K (high thermal conductivity) layer (e.g., a metal layer) is located on the inner surface, outer surface, or inside of one plate of a sample holder (RHC card), only a part of the high-K layer and a part of the sample volume beyond the part of the high-K layer are heated to the desired temperature, while the rest of the high-K layer and the rest of the sample volume are maintained at a much lower temperature during the thermal cycle. Several conditions must be met. The key conditions are: (1) The heat source must directly heat a part of the high-K layer (this part is called the "heating zone", e.g., only this part is directly heated by an LED lamp or has a local electric heater, while the rest is not), (2) The vertical heat transfer between the heating zone and a part of the sample should be much larger than the lateral heat transfer within the high-K material (i.e., in the lateral direction of the high-K material), (3) The relevant sample should have a large lateral-to-vertical dimension ratio, and (4) The heating power of the heating zone must be sufficient to heat the relevant sample volume within a time range where the lateral heat transfer (i.e., heat conduction) is relatively negligible.

[0208] To describe the requirements for meeting condition (2) above, the scaled thermal conductivity (STC ratio) of the vertical heat transferred from the high-K heating zone through the intermediate layer between the high-K and the sample to the sample and the lateral heat transfer within the high-K layer is defined as:

[0209]

[0210] where K k , K s , and K m are the thermal conductivities of the high-K layer, the relevant sample, and the intermediate layer (i.e., the layer between the high-K and the sample), respectively, t k , t s , and t m are the thicknesses of the high-K layer, the sample, and the intermediate layer, respectively; D is the average lateral dimension of the relevant sample, and 0.025 is a scaling factor.

[0211] In some embodiments, to locally heat a part of the high-K layer and a part of the sample volume above this part of the high-K layer to the desired temperature, while maintaining the rest of the high-K layer and the rest of the sample volume at a much lower temperature during the thermal cycle, the scaled thermal conductivity (STM ratio) is 2 or greater, 5 or greater, 10 or greater, 20 or greater, 30 or greater, 40 or greater, 50 or greater, 100 or greater, 1000 or greater, 10000 or greater, 10000 or greater, or within a range between any two values.

[0212] In some preferred embodiments, the scaled thermal conductivity (STM ratio) is in the range of 10 to 20, 30 to 50, 100 to 1000, 1000 to 10000, or 10000 to 1000000.

[0213] To meet the above conditions (2) and (3), in some embodiments, the ratio of the lateral to vertical size (LVS) of the relevant sample is 5, 10, 20, 50, 70, 100, 200, 300, 400, 500, 600, 700, 800, 800, 1,000, 2,000, 5,000, 10,000, 100,000, or within a range between any two values.

[0214] In some preferred embodiments, the LVS ratio of the relevant sample is within the range of 5 to 10, 10 to 50, 50 to 100, 100 to 500, 500 to 1,000, 1,000 to 10,000, or 10,000 to 100,000.

[0215] In certain embodiments, the thickness of the relevant sample is reduced (which also contributes to the sample heating rate), and the thickness of the relevant sample is 0.05 μm, 0.1 μm, 0.2 μm, 0.5 μm, 1 μm, 2 μm, 5 μm, 10 μm, 20 μm, 30 μm, 40 μm, 50 μm, 60 μm, 70 μm, 80 μm, 90 μm, 100 μm, 200 μm, 300 μm, or within a range between any two values.

[0216] In some preferred embodiments, the thickness of the relevant sample is 0.05 μm and 0.5 μm, 0.5 μm and 1 μm, 1 μm and 5 μm, 5 μm and 10 μm, 10 μm and 30 μm, 30 μm and 50 μm, 50 μm and 70 μm, 70 μm and 100 μm, 100 μm and 200 μm, or 200 μm and 300 μm.

[0217] C. Large sample to non-sample thermal mass ratio (NSTM ratio)

[0218] An increase in the sample to non-sample thermal mass ratio can shorten the heating time, reduce the heating energy, and increase the energy efficiency. In embodiments where the sample is sandwiched between two plates, assuming no heat loss in these volumes, the thermal mass ratio can be estimated by considering only the volume of the relevant sample and the portions of the two plates that sandwich the relevant sample. Thus, one parameter for measuring the thermal mass ratio is the ratio of the "specific surface area thermal mass" of the relevant sample to the non-sample (the portion of the plates that sandwich the relevant sample along with the heating / cooling layers on the relevant sample and the plate portions). The term "specific surface area thermal mass" of a material refers to the volumetric specific heat of the material multiplied by its thickness.

[0219] Therefore, assuming that heat losses due to heat conduction and radiation can be neglected, the sample to non-sample thermal mass ratio is the ratio of the useful thermal energy (which directly heats the relevant sample) to the "wasted thermal energy" (which heats the non-sample material).

[0220] For example, the volumetric specific heat of water is 4.2 J / (cm 3 -C), so the areal specific heat of a 30-μm thick water layer is 1.26×10 -2 J / (cm 2 -C). The volumetric specific heat of PMMA is 1.77 J / (cm 3 -C), so the areal specific heat of a 25-μm thick PMMA layer is 4.43×10 -3 J / (cm 2 -C), which is ~2.8 times smaller than the areal specific heat of the 30-μm water layer. Gold has a volumetric specific heat of 2.5 J / (cm 3 -C), so the areal specific heat of a 0.5-μm thick gold layer is 1.25×10 -4 J / (cm 2 -C), which is 100 times smaller than the areal specific heat of the 30-μm water layer and can be neglected. The negligible areal specific heat of Au is due to its thin thickness.

[0221] If, in an RHC card embodiment, the relevant sample is sandwiched between two plates each 25 μm thick and the heating / cooling layer is 0.5 μm thick, then in this case the thermal mass ratio of sample to non-sample is 1.4. That is, when heat losses due to heat conduction and radiation are neglected, the ratio of useful energy to wasted energy is 1.4, and the ratio of useful energy to total heating energy is 58%.

[0222] In some embodiments, the thermal mass ratio of sample to non-sample (NSTM ratio) is 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 1, 1.5, 2, 3, 4, 5, 10, 20, 30, 40, 50, 60, 70, 100, 200, 300, 1000, 4000, or in the range between any two values.

[0223] In preferred embodiments, the thermal mass ratio of sample to non-sample (NSTM ratio) is in the range of 0.1 to 0.2, 0.2 to 0.5, 0.5 to 0.7, 0.7 to 1, 1 to 1.5, 1.5 to 5, 5 to 10, 10 to 30, 30 to 50, 50 to 100, 100 to 300, 300 to 1000, or 1000 to 4000.

[0224] To make the thermal mass ratio of sample to non-sample high, it is necessary to keep the thermal mass of the non-sample region low, which in turn requires the plates and the heating / cooling layer to be thin, and / or have a low volumetric specific heat.

[0225] To achieve a large thermal mass ratio, in one embodiment, a thin material with multiple layers or a hybrid material is used. For example, the thickness of a carbon fiber layer with a plastic sheet or carbon mixed with plastic can be 0.1μm, 0.2μm, 0.5μm, 1μm, 2μm, 5μm, 10μm, 25μm, 50μm, or within the range between any two values.

[0226] D. Thin thickness of related samples and large lateral / vertical size ratio (LVS ratio)

[0227] The term "lateral to vertical size ratio of a sample" or "LVS ratio of a sample" refers to the ratio of the average lateral size to the average vertical size of the volume of the related sample. In embodiments where heating and / or cooling mainly come from the vertical direction, a larger LVS ratio of the sample can reduce wasted heating energy and increase the heating rate and / or cooling rate, and can reduce lateral heat conduction losses at the edges of the related sample relative to the total thermal energy. All of these can increase and / or can increase the cooling time.

[0228] In some embodiments, the LVS ratio of the related sample is 5, 10, 20, 50, 70, 100, 200, 300, 400, 500, 600, 700, 800, 800, 1,000, 2000, 5000, 10,000, 100,000, or within the range between any two values.

[0229] In some preferred embodiments, the LVS ratio of the related sample is within the range of 5 to 10, 10 to 50, 50 to 100, 100 to 500, 500 to 1,000, 1000 to 10,000, or 10,000 to 100,000.

[0230] For example, the lateral size of the sample is 15mm and the thickness is 30μm, so the LVS of the sample is 500.

[0231] In certain embodiments, the thickness of the related sample is reduced (which also helps the heating rate of the sample), and the thickness of the related sample is 0.05μm, 0.1μm, 0.2μm, 0.5μm, 1μm, 2μm, 5μm, 10μm, 20μm, 30μm, 40μm, 50μm, 60μm, 70μm, 80μm, 90μm, 100μm, 200μm, 300μm, or within the range between any two values.

[0232] In some preferred embodiments, the thickness of the relevant sample is 0.05 μm and 0.5 μm, 0.5 μm and 1 μm, 1 μm and 5 μm, 5 μm and 10 μm, 10 μm and 30 μm, 30 μm and 50 μm, 50 μm and 70 μm, 70 μm and 100 μm, 100 μm and 200 μm, or 200 μm and 300 μm.

[0233] E. Thin thickness of non-sample and large lateral / vertical size ratio (LVS ratio)

[0234] The term "lateral to vertical size ratio of non-sample" or "LVS ratio of non-sample" refers to the ratio of the average lateral size of the portions of the two plates sandwiching the relevant sample (which is the same as the average lateral size of the relevant sample volume) to its thickness. A large LVS ratio of the non-sample can reduce the lateral heat conduction loss at the edges of the non-sample relative to the total thermal energy.

[0235] In some embodiments, the LVS ratio of the non-sample is 5, 10, 20, 50, 70, 100, 200, 300, 400, 500, 600, 700, 800, 800, 1,000, 2,000, 5,000, 10,000, 100,000, or within a range between any two values.

[0236] In preferred embodiments, the LVS ratio of the non-sample is in the range of 5 to 10, 10 to 50, 50 to 100, 100 to 500, 500 to 1,000, 1,000 to 10,000, or 10,000 to 100,000.

[0237] For example, two plates with a thickness of 25 μm sandwich a sample with a lateral size of 5 mm or greater of the relevant sample, so the LVS of the non-sample of each plate is 200 or higher.

[0238] To shorten the heating time, reduce the heating energy, and improve the energy efficiency, the lateral heat conduction through the non-sample material (on the sample holder) should be reduced.

[0239] In particular, when the first plate and the second plate are made of materials that are not good thermal materials, the thickness of the plates should be minimized.

[0240] In some embodiments, the thickness of each of the first plate, the second plate, or both plates is 10 nm, 100 nm, 200 nm, 300 nm, 400 nm, 500 nm, 1 μm, 2.5 μm, 5 μm, 10 μm, 25 μm, 50 μm, 100 μm, 200 μm, or 500 μm, 1,000 μm, or within a range between any two values.

[0241] In some preferred embodiments, the thickness of the first plate, or the second plate, or each of the two plates is 10 nm, 100 nm, 200 nm, 300 nm, 400 nm, 500 nm, 1 μm, 2.5 μm, 5 μm, 10 μm, 25 μm, 50 μm, 75 μm, or within a range between any two values.

[0242] The first and second plates may have the same thickness or different thicknesses, and may be made of the same material or different materials.

[0243] In some preferred embodiments, the thickness of the first plate, or the second plate, or each of the two plates is between 10 nm and 500 nm, between 500 nm and 1 μm, between 1 μm and 2.5 μm, between 2.5 μm and 5 μm, between 5 μm and 10 μm, between 10 μm and 25 μm, between 25 μm and 50 μm, between 50 μm and 100 μm, between 100 μm and 200 μm, or between 200 μm and 500 μm, or between 500 μm and 1000 μm.

[0244] In some preferred embodiments, the first and second plates are plastic, thin glass, or materials with similar physical properties. The thickness of the first plate or the second plate is 100 nm, 500 nm, 1 μm, 5 μm, 10 μm, 25 μm, 50 μm, 100 μm, 175 μm, 250 μm, or within a range between any two values.

[0245] In some preferred embodiments, the first and second plates are plastic, thin glass, or materials with similar physical properties. The thickness of the first plate is 5 μm, 10 μm, 25 μm, 50 μm, or within a range between any two values; while the thickness of the second plate (the plate with the heating or cooling layer) is 100 nm, 500 nm, 1 μm, 5 μm, 10 μm, or within a range between any two values.

[0246] F. Cooling layer with high K and / or high heat capacity ratio (KC ratio).

[0247] Since any heat conduction through non-sample materials will waste energy, and since lateral heat conduction has a much longer heat path than vertical heat conduction, the energy wasted in lateral heat conduction in non-sample materials should be minimized. One way to minimize this wasted energy is to use a high thermal conductivity (high K) or more precisely a material with a high ratio of thermal conductivity to capacity (KC ratio) as the cooling layer. For a given thermal conductivity, a given temperature change, and a given geometry, high K and / or high KC ratio materials will require much less energy to heat than low K and / or low KC ratio materials.

[0248] In some embodiments, the KC ratio of the material used for the cooling layer is equal to or higher than 0.1 cm 2 / sec, 0.2 cm 2 / sec, 0.3 cm 2 / sec, 0.4 cm 2 / sec, 0.5 cm 2 / sec, 0.6 cm 2 / sec, 0.7 cm 2 / sec, 0.8 cm 2 / sec, 0.9 cm 2 / sec, 1 cm 2 / sec, 1.1 cm 2 / sec, 1.2 cm 2 / sec, 1.3 cm 2 / sec, 1.4 cm 2 / sec, 1.5 cm 2 / sec, 1.6 cm 2 / sec, 2 cm 2 / sec, 3 cm 2 / sec, or within a range between any two values.

[0249] In some preferred embodiments, the KC ratio of the cooling layer is between 0.5 cm 2 / sec and 0.7 cm 2 / sec, 0.7 cm 2 / sec and 0.9 cm 2 / sec, 0.9 cm 2 / sec and 1 cm 2 / sec, 1 cm 2 / sec and 1.1 cm 2 / sec, 1.1 cm 2 / sec and 1.3 cm 2 / sec, 1.3 cm 2 / sec and 1.6 cm 2 / sec and 1.6 cm.

[0250] In some embodiments, a high thermal conductivity (i.e., high K) material is used for the cooling layer. And the thermal conductivity of the high K material is equal to or greater than 50 W / (m·K), 80 W / (m·K), 100 W / (m·K), 150 W / (m·K), 200 W / (m·K), 250 W / (m·K), 300 W / (m·K), 350 W / (m·K), 400 W / (m·K), 450 W / (m·K), 500 W / (m·K), 600 W / (m·K), 1000 W / (m·K), 5000 W / (m·K), or within a range between any two values.

[0251] In some preferred embodiments, a high thermal conductivity (i.e., high-K) material is used for the cooling layer, and the thermal conductivity of the high-K material is in the range of 50 W / (m·K) to 100 W / (m·K), 110 W / (m·K) to 200 W / (m·K), 200 W / (m·K) to 400 W / (m·K), 400 W / (m·K) to 600 W / (m·K), or 400 W / (m·K) to 5000 W / (m·K).

[0252] In some embodiments, the high-K material is selected from metals, semiconductors, and permits a thermal conductivity higher than 50 W / (m·K), as well as any combination (including any mixture). In some embodiments, the high-K material is selected from gold, copper, silver, and aluminum, as well as any combination (including any mixture). In some embodiments, the high-K material is selected from carbon particles, carbon nanotubes, graphite, silicon, and any combination (including any mixture).

[0253] G-1. The area of the cooling zone is greater than the area of the laterally-related sample area and the heating zone area

[0254] To effectively cool the sample while reducing wasted energy in non-sample materials, in some embodiments, a high-K and / or high-KC ratio material (referred to as "high-K material") is used as the main channel for removing heat from the sample. The area of the high-K cooling zone (layer) should be greater than the relevant lateral dimension of the sample.

[0255] In certain embodiments, the area of the cooling zone (layer) is 1.5, 2, 3, 4, 5, 10, 20, 50, 70, 100, 200, 300, 400, 500, 600, 700, 800, 800, 1,000, 2,000, 5,000, 10,000, 100,000 times greater than the lateral area of the relevant sample, or within the range between any two values.

[0256] In preferred embodiments, the area of the cooling zone (layer) is 1.5 to 5, 5 to 10, 10 to 50, 50 to 100, 100 to 500, 500 to 1,000, 1,000 to 10,000, or 10,000 to 100,000 times greater than the lateral area of the relevant sample.

[0257] To increase the cooling rate and thermal cycling efficiency, in certain embodiments, the area of the high-K cooling layer (zone) should be greater than the area of the heating zone.

[0258] In some embodiments, the area of the cooling zone (layer) is 1.1, 1.5, 2, 3, 4, 5, 10, 20, 30, 40, 50, 70, 100, 200, 300, 400, 500, 600, 700, 800, 800, 1,000, 5,000, 10,000, 100,000 times larger than the area of the heating zone (layer), or within a range between any two values.

[0259] In preferred embodiments, the area of the cooling zone (layer) is 1.1 to 1.5, 1.5 to 5, 5 to 10, 10 to 50, 50 to 100, 100 to 500, 500 to 1,000, 1,000 to 10,000, or 10,000 to 100,000 times larger than the lateral area of the heating zone (layer).

[0260] G-2. The cooling zone area and the heating zone area are the same as the laterally related sample area

[0261] In certain embodiments, the cooling zone area and the heating zone area are the same as the laterally related sample area, the laterally related sample area is much smaller than the total sample area on the plate, and is smaller than the area of the plate. The area of the cooling zone is 1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 、1 mm 2 ,

[0262] The cooling zone can have different shapes. In certain embodiments, there is more than one cooling zone on a plate, and the cooling zones are separated from each other by a low thermal conductivity material (such as air or plastic).

[0263] H. Heating zone with high K and / or high ratio of thermal conductivity to capacity (KC ratio)

[0264] Since any heat conduction of wasted energy and non-sample materials in lateral heat conduction has a much longer heat path than vertical heat conduction, the energy wasted in lateral heat conduction in non-sample materials should be minimized. One way to minimize this wasted energy is to use a material with a ratio of thermal conductivity to capacity (KC) in the heating zone, which will require much less thermal energy for a given thermal conductivity, a given temperature change, and a given geometry.

[0265] In some embodiments, the KC ratio material for the heating layer is equal to or higher than 0.1 cm 2 / sec, 0.2 cm 2 / sec, 0.3 cm 2 / sec, 0.4 cm 2 / sec, 0.5 cm 2 / sec, 0.6 cm 2 / sec, 0.7 cm 2 / sec, 0.8 cm 2 / sec, 0.9 cm 2 / sec, 1 cm 2 / sec, 1.1 cm 2 / sec, 1.2 cm 2 / sec, 1.3 cm 2 / sec, 1.4 cm 2 / sec, 1.5 cm 2 / sec, 1.6 cm 2 / sec, 2 cm 2 / sec, 3 cm 2 / sec, or within a range between any two values.

[0266] In some preferred embodiments, the KC ratio of the heating layer is between 0.5 cm 2 / sec and 0.7 cm 2 / sec, 0.7 cm 2 / sec and 0.9 cm 2 / sec, 0.9 cm 2 / sec and 1 cm 2 / sec, 1 cm 2 / sec and 1.1 cm 2 / sec, 1.1 cm 2 / sec and 1.3 cm 2 / sec, 1.3 cm 2 / sec and 1.6 cm 2 / sec, 1.6 cm 2 / sec and 2 cm 2 / sec, or 2 cm 2 / sec and 3 cm 2 / sec.

[0267] In some embodiments, a high thermal conductivity (i.e., high-K) material is used for the heating layer, and the thermal conductivity of the high-K material is equal to or greater than 50 W / (m·K), 80 W / (m·K), 100 W / (m·K), 150 W / (m·K), 200 W / (m·K), 250 W / (m·K), 300 W / (m·K), 350 W / (m·K), 400 W / (m·K), 450 W / (m·K), 500 W / (m·K), 600 W / (m·K), 1000 W / (m·K), 5000 W / (m·K), or within a range between any two values.

[0268] In some preferred embodiments, a high thermal conductivity (i.e., high-K) material is used for the heating layer, and the thermal conductivity of the high-K material is in the range of 50 W / (m·K) to 100 W / (m·K), 110 W / (m·K) to 200 W / (m·K), 200 W / (m·K) to 400 W / (m·K), 400 W / (m·K) to 600 W / (m·K), or 400 W / (m·K) to 5000 W / (m·K).

[0269] In some embodiments, the high-K material is selected from metals, semiconductors, and allows a thermal conductivity higher than 50 W / (m·K), and any combination (including any mixture). In some embodiments, the high-K material is selected from gold, copper, silver, and aluminum, and any combination (including any mixture). In some embodiments, the high-K material is selected from carbon particles, carbon nanotubes, graphite, silicon, and any combination (including any mixture).

[0270] To receive light energy through the heating zone (layer), a surface with enhanced thermal radiation will be used (on one or both sides of the heating zone). The surface with enhanced thermal radiation absorption can be achieved by directly changing the surface structure (e.g., patterning nanostructures), coating with a high thermal radiation material (e.g., coating with black paint), or both.

[0271] The surface with enhanced thermal radiation has a high average light absorption rate (e.g., the black paint used in our experiments). In certain embodiments, the average light absorption rate of the surface of the heating zone is 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 100%, or within a range between any two values.

[0272] In certain preferred embodiments, the average light absorption rate of the surface of the heating zone is in the range of 30% to 40%, 40% to 60%, 60% to 80% to 90%, or 90% to 100%.

[0273] In some preferred embodiments, the average light absorption rate of the surface of the heating zone is in the range of 30% to 100%, 50% to 100%, 70% to 100%, or 80% to 100%.

[0274] In some embodiments, the value of the average optical absorptance of the surface of the heating zone is obtained by taking the average over the wavelength ranges of 400 nm to 800 nm, 700 nm to 1500 nm, 900 nm to 2000 nm, or 2000 nm to 20000 nm.

[0275] Increased radiative cooling

[0276] In some embodiments, rapid temperature cycling is achieved by increasing the percentage of radiative cooling in the total cooling of the sample and the sample holder during the thermal cycle (i.e., transferring heat to the environment), preferably by using a material with high thermal conductivity as the material for radiative cooling. One reason is that cooling by lateral heat conduction requires heating a variety of non-sample materials and wastes energy. Another reason is that radiative cooling is proportional to the fourth power of the temperature and can be more efficient than heat conduction in thin films.

[0277] To enhance radiative cooling, in some embodiments, radiative cooling uses a cooling layer (cooling zone) that is enhanced for radiative cooling. This enhancement includes (i) increasing the thermal conductivity of the cooling zone (layer), (ii) expanding the area of the cooling zone (layer), (iii) enhancing the surface thermal radiation of the cooling zone, and (iv) combinations thereof.

[0278] Examples of materials with high thermal conductivity are metals (such as gold, silver, CoBr, aluminum), semimetals, semiconductors (such as silicon), or combinations thereof.

[0279] To further enhance the thermal radiation of the cooling zone (layer), a surface enhanced for thermal radiation will be used (on one or both sides of the cooling zone). The surface enhanced for thermal radiation can be achieved by directly changing the structure of the surface (e.g., patterning nanostructures), coating with a material with high thermal radiation (e.g., coating with black paint), or both.

[0280] The surface enhanced for thermal radiation has a high average optical absorptance (e.g., the black paint used in our experiments). In some embodiments, the average optical absorptance of the surface of the cooling zone is 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 100%, or in the range between any two values.

[0281] In some preferred embodiments, the average optical absorptance of the surface of the cooling zone is in the range of 30% to 40%, 40% to 60%, 60% to 80% to 90%, or 90% to 100%.

[0282] In some preferred embodiments, the average optical absorptance of the surface of the cooling zone is in the range of 30% to 100%, 50% to 100%, 70% to 100%, or 80% to 100%.

[0283] In some embodiments, the value of the average light absorption rate of the surface of the cooling zone is obtained by taking the average of the wavelength ranges of 400 nm to 800 nm, 700 nm to 1500 nm, 900 nm to 2000 nm, or 2000 nm to 20000 nm.

[0284] In some embodiments, the surface heat radiation enhancement layer is a black coating, a plasma structure, a nanostructure, or any combination thereof.

[0285] The high heat radiation material is a polymer mixture that appears black to the human eye (commonly referred to as a "black coating"). The high heat radiation material includes, but is not limited to, a mixture of a polymer and nanoparticles. An example of the nanoparticles is a mixture of black carbon nanoparticles, carbon, nanotubes, graphite particles, graphene, metal nanoparticles, semiconductor nanoparticles, or a combination thereof.

[0286] The high heat radiation material also includes a material that is deposited or formed on the surface of the layer and appears black to the human eye. The material includes, but is not limited to, carbon black nanoparticles, carbon, nanotubes, graphite particles, graphene, metal nanoparticles, semiconductor nanoparticles, or a combination thereof.

[0287] The plasma structure includes a nanostructured plasma structure.

[0288] In some embodiments, the cooling layer includes a high thermal conductivity metal (50 W / (m·K) or higher) layer having a surface heat radiation enhancement layer. In some embodiments, the surface heat radiation enhancement layer has a low lateral thermal conductivity, which is attributed to an ultrathin layer, a low thermal conductivity, or both.

[0289] Thermal radiation cooling percentage.

[0290] In some embodiments, the thermal radiation cooling is achieved by increasing the area of the radiation cooling layer (i.e., a high-K material, unless otherwise specified), and the area of the radiation cooling layer is 1.2, 1.5, 2, 3, 4, 5, 10, 20, 30, 40, 50, 60, 70, 80, 100, 200, 300, 400, 500, 600, 700, 800, 800, 1,000, 2,000, 5,000, 10,000, 100,000 times, or a multiple within the range between any two values, larger than the lateral area of the relevant sample.

[0291] In a preferred embodiment, the area of the radiation cooling zone (layer) is 1.2 to 3, 3 to 5, 5 to 10, 10 to 50, 50 to 100, 100 to 500, 500 to 1,000, 1,000 to 10,000, or 10,000 to 100,000 times larger than the lateral area of the relevant sample.

[0292] In some embodiments, during the thermal cycling process, the ratio of the radiative cooling by the cooling zone (layer) to the total cooling of the sample and the sample holder is 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99%, or within a range between any two values.

[0293] In some preferred embodiments, during the thermal cycling process, the ratio of the radiative cooling of the cooling zone (layer) to the total cooling of the sample and the sample holder is within a range between 10% and 20%, 20% and 30%, 30% and 40%, 40% and 50%, 50% and 60%, 60% and 70%, 70% and 80%, 80% and 90%, or 90% and 99%.

[0294] J. Cooling Layer Thickness Control

[0295] In certain embodiments, the thickness of the cooling layer is configured to promote local optimization of heating and / or optimization of energy efficiency. If the cooling zone (layer) is too thick, a larger percentage of the heating energy will be wasted by the cooling layer, prolonging the heating time (for a given heating power). On the other hand, if the cooling zone is too thin, the cooling time will be significantly longer. Therefore, the cooling layer thickness should be optimized for rapid heating and cooling.

[0296] It has been found experimentally that the thickness of the high-K cooling layer can regulate the cooling rate. By selecting an appropriate high-K cooling layer thickness and an appropriate LED power density, rapid heating and cooling can be achieved.

[0297] Since the thermal conductivity of the layer is proportional to the thermal conductivity of the material multiplied by the layer thickness, the product should be optimized.

[0298] In some embodiments, the thermal conductivity of the cooling zone (layer) multiplied by its thickness is 6×10 -5 W / K, 9×10 -5 W / K, 1.2×10 -4 W / K, 1.5×10 -4 W / K, 1.8×10 -4 W / K, 2.1×10 -4 W / K, 2.7×10 -4 W / K, 3×10 -4 W / K, 1.5×10 - 4 W / K, or within a range between any two values.

[0299] In some preferred embodiments, the thermal conductivity of the cooling zone (layer) multiplied by its thickness is between 6×10 -5 W / K and 9×10 -5 W / K, 9×10 -5 W / K and 1.5×10-4 W / K, 1.5×10 -4 W / K to 2.1×10 -4 W / K, 2.1×10 -4 W / K to 2.7×10 -4 W / K, 2.7×10 -4 W / K to 3×10 -4 W / K, or 3×10 -4 W / K to 1.5×10 -4 W / K within the range.

[0300] In some preferred embodiments, the thermal conductivity of the cooling zone (layer) multiplied by its thickness is in the range of 9×10 -5 W / K to 2.7×10 - 4 W / K, 9×10 -5 W / K to 2.4×10 -4 W / K, 9×10 -5 W / K to 2.1×10 -4 W / K or 9×10 -5 W / K to 1.8×10 -4 W / K within the range.

[0301] In one embodiment, the cooling zone comprises a layer of gold material having a thickness in the range of 200 nm to 800 nm. In another embodiment, the cooling zone comprises a layer of gold material having a thickness in the range of 300 nm to 700 nm.

[0302] K. The large conductance between the sample and the heating or cooling zone

[0303] To rapidly heat and cool the sample, the heat conduction per unit area between the relevant sample and the heating layer and / or cooling layer should be large. The heat conduction per unit area is equal to the conductivity (per unit volume) divided by the material thickness of the material between the HC layer and the sample. For example, for a 100 nm thick PS as the second plate having an HC layer on one surface and a sample on the other surface, the conductivity between the HC layer and the sample is -1000 W / (m 2 ·K).

[0304] Based on experiments, in some embodiments of the RHC card, the material between the heating zone and the relevant sample has a thermal conductivity and thickness configured to be approximately 1000 W / (m 2 ·K) or higher.

[0305] In some embodiments of the RHC card, the material between the heating zone and the relevant sample has a thermal conductivity and thickness, and the thickness is configured to have a value equal to or greater than 1000 W / (m 2 ·K), 2000 W / (m 2·K), 3000 W / (m 2 ·K), 4000 W / (m 2 ·K), 5000 W / (m 2 ·K), 7000 W / (m 2 ·K) Km 2 ·K), 10000 W / (m 2 ·K), 20000 W / (m 2 ·K), 50000 W / (m 2 ·K), 50000 W / (m 2 ·K), 100000 W / (m 2 ·K), or the conductivity per unit area within the range of any of these values.

[0306] The preferred conductivity per unit area of the material between the heating zone and the associated sample is 1000 W / (m 2 ·K) to 2000 W / (m 2 ·K), 2000 W / (m 2 ·K) to 4000 W / (m 2 ·K), 4000 W / (m 2 ·K) to 10000 W / (m 2 ·K), or 10000 W / (m 2 ·K) to 100000 W / (m 2 ·K).

[0307] In another preferred embodiment, the distance between the heating zone and the associated sample is zero, so the conductance per unit area of the material between the heating zone and the associated sample is infinite.

[0308] In some embodiments, the heating layer or the cooling layer is separated from the associated sample by a thin plastic sheet (or film) with a thermal conductivity in the range of 0.1 W / (m·K) to 0.3 W / (m·K), and the thickness of the thin plastic layer is 0 nm, 10 nm, 50 nm, 100 nm, 200 nm, 300 nm, 400 nm, 500 nm, 1 μm, 2.5 μm, 5 μm, 10 μm, 25 μm, 50 μm, 75 nm, 100 μm, 150 μm, or within the range between any two values.

[0309] In some preferred embodiments, the thickness of the thin plastic sheet (or film) separating the associated sample from the heating layer or the cooling layer is 0 nm to 100 nm, 100 nm to 500 nm, 500 nm to 1 μm, 1 μm to 5 μm, 5 μm to 10 μm, 10 μm to 25 μm, 25 μm to 50 μm, 50 μm to 75 μm, 75 μm to 100 μm, or 100 μm to 150 μm.

[0310] In a preferred embodiment of the RHC card, the thickness of the thin plastic sheet (or film) that separates the relevant sample from the heating or cooling layer is 1 nm, 10 nm, 0.1 μm, 0.5 μm, 1 μm, 5 μm, 10 μm, 20 μm, 25 μm, or within a range between any two values.

[0311] L. Smaller lateral diffusion of the relative reagent

[0312] To make the biochemical reactions in the relevant sample volume substantially uniform during temperature changes or thermal cycling, the average lateral area of the relevant sample should be significantly larger than the lateral diffusion of the nucleic acid and / or other reagents used for molecular amplification and / or reaction. In this way, during temperature changes or thermal cycling, most of the molecules within the relevant sample volume do not have enough time to diffuse out of the relevant sample volume, and most of the molecules outside the relevant sample volume do not have enough time to diffuse into the relevant sample volume.

[0313] Considering that for molecules with a molecular weight of approximately 600 Da, the thermal cycling duration is 3 minutes, the diffusion constant is -1×10 - 6 cm 2 / s, and the diffusion length is -130 μm.

[0314] In certain embodiments, during thermal cycling or reaction, the ratio of the average lateral dimension of the relevant sample volume to the diffusion length of the reagent is equal to or greater than 5, 6, 7, 10, 15, 20, 25, 30, 40, 50, 60, 70, 80, 90, 100, 150, 200, 500, 1000, 5000, 10000, 100000, or within a range between any two values.

[0315] In some preferred embodiments, during thermal cycling or reaction, the ratio of the average lateral dimension of the relevant sample volume to the diffusion length of the reagent is within the range of 5 to 10, 10 to 30, 30 to 60, 6 to 100, 100 to 200, 200 to 500, 500 to 1000, 1000 to 5000, 5000 to 10000, or 10000 to 100000.

[0316] In some preferred embodiments, during thermal cycling or reaction, the ratio of the average lateral dimension of the relevant sample volume to the diffusion length of the reagent is within the range of 5 to 10, 10 to 30, 30 to 60, 6 to 100, 100 to 200, 200 to 500, 500 to 1000, 1000 to 5000, 5000 to 10000, or 10000 to 100000.

[0317] In certain preferred embodiments, the average lateral dimension of the relevant volume is 1 mm, 2 mm, 3 mm, 5 mm, 6 mm, 7 mm, 8 mm, 9 mm, 10 mm, 12 mm, 15 mm, 20 mm, 30 mm, 40 mm, 50 mm, 70 mm, 100 mm, 200 mm, or within a range between any two of these values.

[0318] In some preferred embodiments, the average lateral dimension of the relevant volume is in the range of 1 mm to 5 mm, 5 mm to 10 mm, 10 mm to 20 mm, 20 mm to 40 mm, 40 mm to 70 mm, 70 mm to 100 mm, or 100 mm to 200 mm.

[0319] In another preferred embodiment, the average lateral dimension of the relevant volume is in the range of 1 mm to 5 mm, 1 mm to 10 mm, or 5 mm to 20 mm.

[0320] M. No Edge Sealing or Simple Edge Sealing

[0321] To simplify the operation and cost of the sample holder, in certain embodiments, there is no seal between the two plates defining the sample; that is, the sample sandwiched between the plates can evaporate into the environment from the sample edges. However, in our experiments, we found that in our sample card configuration, due to the large ratio of the lateral sample area to the sample edge area, this evaporation is negligible relative to the total sample volume; the plates prevent most of the evaporation.

[0322] In some embodiments, a closed-loop spacer or some discontinuous spacer walls can be placed on one or both plates to reduce or eliminate sample evaporation.

[0323] P-2 Forced Air Cooling

[0324] In certain embodiments, a forced air cooling / circulation system is present near the RHC card to accelerate the cooling process. Examples of forced air cooling systems include, but are not limited to, a fan that circulates cold air near the card, a plurality of fans that circulate cold air near the card, a cooling source that cools the air near the card, a cooling pad that directly contacts the card, or a combination thereof.

[0325] In certain embodiments, there is a forced air cooling / circulation system that cools the air on the top surface of the card.

[0326] In certain embodiments, there is a forced air cooling / circulation system that cools the air on the bottom surface of the card.

[0327] In certain embodiments, there is a forced air cooling / circulation system that cools the air around all surfaces of the card.

[0328] 2. Mechanical Structure Design

[0329] N. Movable Plate and Extrusion Open Flow, Hinge, Open Notch, Groove Edge and Slide Block

[0330] For simple sample loading, in certain embodiments of the present invention, the two plates of the RHC card can move relative to each other into different configurations. The sample is deposited in the open configuration of the plates, and then the plates are pressed into a closed configuration. During the pressing process, the sample will flow into a thin layer between the plates, and this flow is called "extrusion open flow" because there is sufficient space between the plates to allow the sample to flow.

[0331] In certain embodiments, a space for adjusting the sample thickness is added to one or both plates, so the device for quickly changing the temperature of a fluid sample comprises:

[0332] A first plate (10), a second plate (20), a heating layer (112-1) and a cooling layer (112-2), wherein:

[0333] The first plate and the second plate can move relative to each other into different configurations;

[0334] Each of the first plate and the second plate has a sample contact area on its respective inner surface for contacting the fluid sample; wherein the sample contact areas face each other, are separated by an average spacing of 200 μm or less, and are capable of clamping the sample between them;

[0335] Heating layer:

[0336] Is positioned on the inner surface, outer surface or inside of one of the plates,

[0337] Is configured to heat a relevant volume of the sample, wherein the relevant volume of the sample is part or all of the sample being heated to a desired temperature; and

[0338] Cooling layer:

[0339] Is located on the inner surface, outer surface or inside of one of the plates;

[0340] Is configured to cool the relevant sample volume; and

[0341] Comprises a material layer with a thermal conductivity to heat capacity ratio of 0.6 cm 2 / sec or greater;

[0342] One of the configurations is an open configuration, wherein: the two plates are partially or completely separated and the average spacing between the plates is at least 300 μm;

[0343] Another of these configurations is a closed configuration that is configured after depositing a fluid sample on one or both sample contact regions of the open configuration; and in the closed configuration: at least a portion of the sample is defined into a layer by two plates, where the average sample thickness is 200 μm or less; and

[0344] In some embodiments, the heating layer and the cooling layer are the same material layer with heating and cooling zones, and where the heating and cooling zones may have the same area or different areas.

[0345] In some embodiments, a sample holder with a movable plate (also referred to as an "RHC card" or "Q card") further includes hinges, notches, grooves that facilitate the manipulation of the sample holder and the measurement of the sample. Additionally, the sample holder can slide into a slider. Hinges, notches, grooves, sliders, and structures for squeezing open fluids, materials, functions, variations, and dimensions are listed, described, and summarized in the PCT applications (designating the United States) No. PCT / US2016 / 045437 filed on August 10, 2016, and No. PCT / US0216 / 051775 filed on September 14, 2016, the U.S. Provisional Application No. 62 / 456065 filed on February 7, 2017, the U.S. Provisional Application No. 62 / 456287 filed on February 8, 2017, and the U.S. Provisional Application No. 62 / 456504 filed on February 8, 2017, all of the entire contents of which are incorporated herein by reference for all purposes.

[0346] Spacer (13)

[0347] In certain embodiments, spacers such as those described in Example SH-5 are used to adjust the sample thickness and make the thickness uniform. Even when both plates are very thin (e.g., 25 μm thick or less), the spacers allow for a uniform sample thickness to be obtained.

[0348] In certain embodiments, the spacers are fixed to one or both plates. In certain embodiments, the spacers are mixed with the sample. In some embodiments, the spacers have a uniform height, and the spacers, together with the first and second plates, adjust the sample layer. In some embodiments, the thickness of the sample layer is substantially equal to the height of the spacers.

[0349] In some embodiments, the plates are flat (e.g., as shown in FIG. 12A). In some embodiments, one or both of the plates include holes (e.g., as shown in FIG. 12B). For example, in certain embodiments, the width of the holes can be less than 500 μm, 200 μm, 100 μm, 50 μm, 25 μm, 10 μm, 5 μm, 2.5 μm, 1 μm, 500 nm, 400 nm, 300 nm, 200 nm, or 100 nm, or within a range between any two values. In certain embodiments, the depth of the holes can be less than 500 μm, 200 μm, 100 μm, 50 μm, 25 μm, 10 μm, 5 μm, 2.5 μm, 1 μm, 500 nm, 400 nm, 300 nm, 200 nm, 100 nm, 50 nm, 20 nm, 10 nm, 5 nm, 2 nm, or 1 nm, or within a range between any two values.

[0350] In some embodiments, one or both of the plates have holes, and most or all of the sample is only within the holes of one plate and is covered by the other plate (not shown in the figure).

[0351] P. Sample cartridge and thermal conduction isolation

[0352] In certain embodiments, the RHC card (sample holder) can be further mounted on the sample cartridge. The cartridge can be configured to slide into or out of a base (also referred to as a "adapter"). The base houses a power supply, a temperature sensor and controller, a signal measurement device, and slots for the sample holder with or without the cartridge to slide into or out of the base.

[0353] In some embodiments, the sample holder, the cartridge (i.e., the sample holder support), or both are "thermally conduction isolated", i.e., they have no or little thermal conduction to the environment during thermal cycling. In this case, cooling during thermal cycling is substantially by thermal radiation (this is referred to as "heat transfer without conduction"). In some embodiments, this "thermal conduction isolation" is achieved in the sample holder, the cartridge, or both by configuring their materials, geometries (including thickness reduction), or both.

[0354] Q. Above combinations

[0355] Embodiments of the RHC card can be any combination of SH-1, SH-2, SH-3, and the specifications described in subsections A to P.

[0356] R. Heating source

[0357] The heating layer or heating / cooling layer in the RHC card is configured to be heated by a heating source, where the heating source transfers thermal energy to the heating / cooling layer by light, electricity, radio frequency (RF) radiation, or a combination thereof.

[0358] S. Base (i.e., adapter)

[0359] In some embodiments, the device further includes a base (adapter) configured to accommodate a sample card, a heat source, a temperature sensor, a part of the overall temperature control (including a smartphone in some embodiments), an additional heat sink (optionally), a fan (optionally), or a combination thereof. In some embodiments, the adapter includes a card slot into which the sample card or sample cassette can be inserted. In some embodiments, after the sample card or sample cassette is fully inserted into the slot, or after reaching a predetermined position in the slot, it is stabilized and held in place without any movement.

[0360] T. Smartphone

[0361] In some embodiments, a smartphone is used to image the sample card, control heating and / or cooling, sense signals, monitor operations using a camera, provide light / energy with a flash, communicate with local or remote devices integrated through the base (adapter) in the system, or a combination thereof.

[0362] U. Applications of Isothermal Nucleic Acid Amplification

[0363] The present invention with minor modifications also provides useful devices and methods for isothermal nucleic acid amplification, where the sample temperature needs to rise from ambient temperature to an elevated temperature (i.e., 65 °C) and be maintained at that temperature for a period of time (i.e., 5 - 10 minutes). In some embodiments, one of the modifications required for isothermal nucleic acid amplification detection is to reduce or eliminate the cooling zone / layer, thereby reducing the heat energy loss from the sample and / or the sample holder to the environment.

[0364] The present invention with minor modifications provides useful devices and methods for reverse transcription polymerase chain reaction, which includes an isothermal process prior to conventional PCR, where the sample temperature needs to rise from ambient temperature to an elevated temperature (i.e., 50 °C) and be maintained at that temperature for a period of time (i.e., 5 - 10 minutes). The present invention with minor modifications provides useful devices and methods for minimizing PCR cross - contamination, such as the method using dUTP and uracil - DNA N - glycosylase, where the sample temperature needs to rise from ambient temperature to an elevated temperature (i.e., 50 °C) and be maintained at that temperature for a period of time (i.e., 1 - 20 minutes).

[0365] Experiments of Certain Embodiments

[0366] Certain embodiments of the present invention have been tested experimentally. Some experimental results are presented herein.

[0367] In some of our experiments, the apparatus shown in Fig. 18A includes a sample holder (e.g., an RHC card), an LED light source (i.e., an energy source) focused by a lens onto a (∼5 mm × 5 mm) area of the sample holder, and a sample holder support made of a thermally insulating material (not shown in Fig. 18A). The sample holder support supports a -2 mm edge at two opposite edges of the second plate (e.g., around the perimeter of the second plate). There is no additional heat sink, and heat is mainly radiated into the open environment (e.g., a room).

[0368] In the experiments described in this section, the sample holder includes a first plate, a second plate, and a heating / cooling layer. One of the plates has spacers. The first and second plates can be moved relative to each other into different configurations; one configuration is an open configuration in which the two plates are separated by an average distance of at least 300 μm. In the open configuration, the sample is deposited on one of the plates. Then another card is placed on top of the sample, and the two plates are pressed together by hand into a closed configuration. In the closed configuration, the spacers adjust the distance between the two plates, and thus the sample thickness is adjusted by the two plates and the spacers. By using appropriate spacers and plates (see other parts of the specification), the sample thickness in the closed configuration can be uniform over a large area and close to the spacer height. It has been found experimentally that the sample thickness is uniform even when the hand pressing force, pressure, and sequence (pressing one area of the RHC card first and then rubbing another area of the RHC card) are different.

[0369] The first plate is made of a poly(methyl methacrylate) (PMMA) film that is 50 μm thick, 20 mm wide, and 20 mm long.

[0370] The second plate is a square that is 20 mm wide and 25 μm thick and is made of a polyethylene terephthalate (PET) film. The second plate has a periodic array of columnar spacers on its inner surface that are 30 μm high, 30 μm × 40 μm in size, and have an 80 μm spacer pitch. The spacers have a uniform height and a flat top surface (note that other types of spacers can be used and will be described later). The spacers fixed to the plate are fabricated by direct imprinting of a flat PMMA plate (other fabrication methods are possible).

[0371] The experiments tested various heating / cooling layers of different materials and geometries on the outer or inner surface of the second plate. An example (shown in FIG. 18A) is that the heating / cooling layer is located on the outer surface of the second plate and covers the entire outer surface of the second plate. The heating / cooling layer includes an Au (gold) film and a black paint layer. One surface of the Au film is in contact with the outer surface of the second plate, while the other surface is coated with black paint. The black paint is a commercial product of a film composed of a mixture of black carbon nanoparticles and a polymer. The average thickness of the black paint is -9 μm (with a thickness variation of -2 μm). The black paint layer can face the incident LED light directly, as shown in FIG. 19. There is a 5-nm Ti adhesion layer between the Au film and the outer surface of the second plate, which improves the adhesion between Au and the second plate. However, the adhesion layer is optional, and due to its thin thickness, it has little or no impact on the thermal performance of the sample holder.

[0372] The heat source can be a blue light-emitting diode (LED) with a central wavelength of 450 nm. As shown in FIG. 19, according to some embodiments, a lens is used to project the light from the LED onto the heating / cooling layer, but only onto the central region of the heating / cooling layer, and generally the size (i.e., area) of the LED light spot on the heating / cooling layer is approximately 5 mm × 5 mm. As shown later, for a given sample card and sample thickness, only the sample at the LED heating point can be changed and / or reach the designed temperature. Therefore, the heating area is approximately 5 mm × 5 mm. In some embodiments, compared with the cooling area of the entire area of the first plate, the cooling area can be approximately 16 times larger than the heating area (i.e., high KC / H ratio = 16).

[0373] The LED heat source is powered by a power supply capable of changing the LED current in less than 100 ms. In some embodiments, an aspherical condenser lens is used to focus the LED light, and the lens has a diameter of 12 mm, a focal length of 10.5 mm, and a numerical aperture (N.A.) of 0.54.

[0374] A temperature-sensitive dye (LDS698) monitors the temperature of the sample in the heating area (i.e., the area directly irradiated by the LED). A photodetector is used to monitor the temperature-sensitive dye and provide feedback to control the LED current and thus control the LED heat source and the temperature of the heating area.

[0375] In the following experiments (Experiments 1 to 12), unless otherwise stated, the sample holder supports the sample by supporting a -2-mm edge at two opposite edges of the second plate, so the sample holder is thermally conductive and isolated from the outside, and the cooling of the sample holder is mainly through thermal radiation cooling. Thermal radiation cooling is mainly provided by the H / C layer because the sample and the plate are poor thermal radiators and have much lower thermal conductivity than the H / C layer. Thermal radiation cooling radiates thermal energy into the open environment (i.e., the room).

[0376] In our experiment, a -5 μL liquid sample with thermal properties close to water was deposited between two plates and approximately in the central region of the plate surface. First, the sample was dropped onto one plate of the RHC card, then another card was placed on top of the sample, and the two plates were manually pressed into a closed configuration. Due to the spacers on the plates, when the two plates were in the closed configuration, the gap between the two plates was adjusted to a 30-μm gap through an array of 30-μm-high spacers, and uniform sample thickness was also found even under different hand pressing forces, pressures, and sequences (first pressing one area and then rubbing into another area of the RHC card). To obtain a good sample thickness by manual pressing, several advantages are provided in the actual use of the present invention.

[0377] For a -5 μL sample between two plates, the sample had a thickness of 30 μm and an area of -166 mm 2 (approximately -13 mm by -13 mm 2 - The lateral shape of the sample was affected by the spacers on the plate, as shown in the top view of FIG. 18B). In some embodiments, the total sample area was more than -6.6 times larger than the heating area (-5 mm × 5 mm). Experiments found that with this setup, only the beverage of the sample above the heating area was heated to the desired temperature. That is, the area (volume) of the heated sample part was approximately 1 / 6 of the total sample area (volume).

[0378] In addition, in this setup, there was no physical wall at the edge of the sample tray, only air. However, as described later, we found that the change in the diameter of the sample tray before and after 30 cycles of PCR was not significant (i.e., almost no difference visible to the naked eye), which means that even without a physical wall (except for the gas-liquid interface) enclosing the liquid sample, sample evaporation could be almost ignored.

[0379] All the spacers used in this experimental section were columns fixed on one plate and having a flat top that could contact the other plate.

[0380] In our experiment, the liquid sample was deposited on one plate, and then the second plate was placed on top of the sample. The plates were pressed together by hand. During the hand pressing, the sample was spread to form a film between the plates. Due to the spacers on the plates (with uniform height), even with hand pressing, the final sample thickness was uniform and was adjusted by the two plate surfaces and the spacer height. In addition, after the sample reached the final thickness and the hand pressing force was removed, the two plates of the sample holder were "self"-held by the capillary force of the liquid sample to "self"-maintain a constant sample thickness. Moreover, even during the thermal cycle at 65 - 95 °C, the capillary force still maintained the sample thickness constant. This self-sampling holding without using any clamps can greatly simplify the device operation and cost.

[0381] Experiment 1

[0382] Light absorption of different H / C layer materials

[0383] In one experiment, the effect of materials used for the H / C layer on the light absorption of LEDs was studied.

[0384] Light absorption spectra of different materials for the heating / cooling (H / C) layer. The optical absorption spectra (i.e., 1-R (light reflection)) of four different H / C layer materials for LED irradiation at 450 nm were experimentally tested: Au (gold) only 500 nm thick (i.e., without black paint), Al (aluminum) only 400 nm thick, Au (500 nm thick) with black paint (9 μm thick), and Al (400 nm thick) with black paint (9 μm thick). We found that, as shown in Figure 20, the black paint coated on Au and Al had a light absorption of -99% over the entire wavelength range from 400 to 800 nm, up to 73% maximum (at a wavelength of -490 nm), and only Au was much smaller after a wavelength of 490 nm; and only for Al, it was 0.1% over the bandwidth from 400 nm to 800 nm. This means that the 9-μm thick black paint used in our experiment greatly enhanced the light absorption and radiation of the H / C layer.

[0385] Experiment 2

[0386] Measurement of the area size of the heating zone

[0387] In another experiment, the area of the heating zone on the HC layer was measured. The experiment found that the vertical heat transfer from the HC layer to the plate and the sample was several orders of magnitude better than the lateral heat conduction in the plate and even in the sample with the HC layer. The area of the heating zone in the sample was approximately the same as the LED irradiation area on the HC layer.

[0388] Experimentally, the sample holder card (shown in Fig. 18A) has a first plate of 50-μm-thick PMMA, a second plate of 25-μm-thick PET, a 30-μm-thick sample notch controlled by spacers, and an H / C layer of gold material on the outer surface of the second plate. The first plate, the second plate, and the Au / black paint HC layer have the same area of 20 mm × 20 mm. The HC layer contains a 500-nm-thick Au (gold) film and a black paint layer. One surface of the gold film is in contact with the outer surface of the second plate, while the other surface is coated with black paint. The black paint is a commercial product of a film composed of a mixture of black carbon nanoparticles and polymers. The average thickness of the black paint is -9 μm (-2-μm thickness variation). The LED heating power projected onto the -5 mm × 5 mm heating zone of the H / C layer is 300 mW. The sample is a 5-μL liquid temperature-sensitive dye LDS698 with a concentration of 2 mg / mL in 60% water and 40% DMSO. The temperature-sensitive dye allows us to optically measure the sample temperature. The 5-μL sample on the RHC card is much larger than the heating zone area, with a thickness of 30 μm and an area of -167 mm 2 The thermal cycle is between 65 °C and 95 °C.

[0389] We have experimentally observed that for a given condition, during the thermal cycle (65 - 95 °C), based on measuring the temperature-sensitive dye, for a sample area of 167 mm 2 , only the sample area (-5 mm × 5 mm) directly irradiated by the LED at the top has a thermal cycle (65 - 95 °C), while the rest of the sample area remains at a nearly constant temperature close to room temperature (i.e., the ambient temperature (e.g., -20 °C)). The thermally cycled area in the sample is approximately 1 / 6 of the total sample area. The transition distance from the thermally cycled area of the sample to the sample area with ambient temperature is measured by the temperature-sensitive dye to be about 2 - 3 mm. This experiment also shows that for a 20 mm × 20 mm area of the Au / black paint HC layer, only the sample area (-5 mm × 5 mm) directly irradiated by the LED is heated. That is, the heating zone is only 1 / 16 of the total HC layer (i.e., high KC / H ratio = 16). As described above, the reason is that in a given RHC card, the vertical heat transfer from the HC layer to the plate and the sample is several orders of magnitude better than the lateral heat conduction in the plate and the sample with the HC layer.

[0390] Experiment 3

[0391] Effect of HC layer area on heating and cooling times

[0392] In another experiment, the effect of the H / C area on heating and cooling times was investigated. Two types of RHC cards were studied.

[0393] The type-1 RHC card uses a disk-shaped HC layer. The type-1 RHC card can include a first plate of 100-μm-thick PMMA (poly(methyl methacrylate)) plate, a second plate of 50-μm-thick PET (polyethylene terephthalate), a 30-μm-thick sample notch controlled by spacers, and an H / C layer of a 700-nm-thick gold film on the outer surface of the second plate. The first plate and the second plate have a square shape and the same area of 20 mm × 20 mm. The second plate has a periodic array of flat-top columnar spacers with a uniform height of 30 μm, a size of 30 μm × 40 μm, and a spacer pitch of 80 μm on its inner surface. The HC layer located at the center of the outer surface of the second plate is a 700-nm-thick Au layer and has a disk shape with different disk diameters for different RHC cards.

[0394] The type-2 RHC card uses a square HC layer. The type-2 RHC card can include a first plate of 50-μm-thick PMMA plate, a second plate of 50-μm-thick PET, a 30-μm-high spacer that controls the sample thickness to 30 μm, and the H / C layer is a 500-nm-thick gold film on the outer surface of the second plate. The first plate has a square shape, an area of 20 mm × 20 mm, and has a periodic array of flat-top columnar spacers with a uniform height of 30 μm, a size of 30 μm × 40 μm, and an 80-μm spacer pitch on its inner surface. The second plate has a square shape and has four different areas for four different HC layers. Two of the second plates have an area of 20 mm × 20 mm for HC layer areas of 10 mm × 10 mm and 20 mm × 20 mm respectively; but the areas of the other two plates, which are 30 mm × 30 mm and 40 mm × 40 mm respectively, are the same as the areas of the HC layers in the HC layer regions.

[0395] When testing the two types of RHC cards, the LED heating power is projected onto the H / C layer with an area of -5 mm × 5 mm to form a heating zone and has a power of 300 mW. The sample is a 5-μL liquid temperature-sensitive dye LDS698 with a concentration of 2 mg / mL in 60% water and 40% DMSO. The temperature-sensitive dye allows us to optically measure the local temperature of the sample. The 5-μL sample on the RHC card, which is much larger than the heating zone area, has a thickness of 30 μm (regulated by the spacers) and an area of -167 mm 2 The thermal cycle is between 65 °C and 95 °C.

[0396] The experimental data (shown in Figures 22 and 23) indicate that as the area of the H / C layer increases, the heating time increases, but the cooling time decreases. For the type-1 RHC card, the HC layer has no direct physical contact with the mechanical support of the card (e.g., the sample holder), so the decrease in the cooling cycle time is mainly due to the increase in the thermal radiation cooling of the HC layer caused by the increase in the radiation cooling area of the HC layer.

[0397] Experiment 4

[0398] Achieve 0.6 s heating, 0.75 s cooling, and 500 mW heating with a 500 nm Au H / C layer

[0399] In another experiment, the heating and cooling cycles of an RHC card (the same as the RHC card shown in FIG. 18A) with a 5 μL water-like sample having a thickness of 30 μm and an LED power of 500 mW were studied. The experimental data shown in FIG. 21 shows 10 cycles between 65°C and 93°C with a heating time of 0.65 seconds (average temperature increase rate of 43°C / second) and a cooling time of 0.75 seconds (average temperature decrease rate of 37°C / second).

[0400] Experiment 5

[0401] (Effect of H / C layer thickness on heating and cooling times)

[0402] In one experiment, the effect of the thickness of the H / C gold layer on heating and cooling times was studied.

[0403] An exemplary RHC card has a first plate of a 100 μm thick PMMA board, a second plate of 50 μm thick PET, an array of 30 μm thick spacers to control the sample thickness, and a gold H / C layer on the outer surface of the second plate. The first plate, the second plate, and the gold H / C layer have the same area of 20 mm × 20 mm. The LED heating power projected onto the -5 mm × 5 mm heating area of the H / C layer is 300 mW. The 5 μL water-like sample on the RHC card is much larger than the heating area, has a thickness of 30 μm and an area of -167 mm 2 The thermal cycle is between 65°C and 95°C.

[0404] The experimental data shown in FIGS. 24A and 24B shows that as the gold thickness of the HC layer varies from 300 nm to 700 nm, the heating time in the thermal cycle slightly increases (from 1.75 seconds to 1.90 seconds), but the cooling time in the thermal cycle decreases with the gold thickness (from 1.5 seconds to 1.3 seconds).

[0405] The cooling cycle time shortens with the gold thickness. This indicates that (a) thermal radiation cooling of the gold HC layer is important in the cooling of the sample, and (b) thermal radiation cooling involves heat conduction from the sample through the gold to the gold surface for radiation. The thicker the gold, the better the heat conduction from the sample to the edge of the gold HC layer.

[0406] The heating cycle time becomes longer as the gold thickness increases. Apparently, thicker gold increases the total heating energy. However, in this experiment, the LED only heats the relevant sample area of -5mm×5mm and the HC layer area of gold material to the maximum cycle temperature, and the thermal mass of gold is small (due to its thin thickness), so the increase in total heating energy is small, resulting in a weakly increasing heating cycle as the gold thickness increases.

[0407] Experiment 6

[0408] (Effect of the distance between the heating / cooling layer and the sample on heating and cooling times)

[0409] In another experiment, the effect of the distance between the HC layer and the sample on heating and cooling times was studied.

[0410] The exemplary RHC card has a first plate of 100μm thick PMMA board, a second plate of PET film with different thicknesses for different RHC cards, a 30μm thick sample thickness controlled by spacers, and the HC layer is made of bare 0.5μm thick gold and is located on the outer surface of the second plate. The first plate, the second plate, and the HC layer of gold material have the same area of 20mm×20mm. The LED heating power projected on the -5mm×5mm heating area of the H / C layer is 300mW. The 5uL water-like sample on the RHC card is much larger than the heating area, with a thickness of 30μm and an area of -167mm 2 The thermal cycle is between 65°C and 95°C.

[0411] The distance between the HC layer and the sample is the distance between the gold surface in contact with the second plate surface and the sample surface in contact with the other second plate surface (i.e., the distance between gold and the sample).

[0412] The sample data shown in Figures 25B and 25C indicate that when the thickness of the second plate changes (thus the distance from gold to the sample) from 25μm to 1000μm, both the heating cycle time and the cooling cycle time increase. However, the heating cycle time increases much more significantly than the cooling cycle time as the second plate thickness increases.

[0413] The data shows that as the thickness of the second plate increases, the energy required to heat and cool the second plate will increase significantly, and the heat conduction between the sample and the HC layer decreases significantly.

[0414] For rapid heating and cooling, the thickness of the second plate (which is physically sandwiched between the sample and the HC layer) should be reduced, and the second plate should be as thin as possible. The preferred thickness of the second plate is 25nm or less. Another preferred thickness of the second plate is 10nm or less.

[0415] Experiment 7

[0416] (Effect of sample thickness on heating and cooling times)

[0417] In another experiment, the effect of the sample thickness sandwiched between two plates on the heating and cooling times was studied.

[0418] The exemplary RHC card has a first plate of a 100-μm-thick PMMA plate, a second plate of a 25-μm-thick PET film, a periodic array of spacers for controlling the sample thickness, and the HC layer is made of bare 0.5-μm-thick gold and is on the outer surface of the second plate. For each different RHC card, the water-like sample has a different notch (i.e., thickness). The first plate, the second plate, and the HC layer of gold material have the same area of 20 mm × 20 mm. The LED heating power projected onto the -5 mm × 5 mm heating zone of the H / C layer is 300 mW. The water-like sample on the RHC card is much larger than the heating zone area, having an area of -167 mm 2 The thermal cycle is between 65 °C and 95 °C.

[0419] Figures 27A and 27B show that when the sample thickness varies from 10 μm to 100 μm, both the heating cycle time and the cooling cycle time increase. However, the heating cycle time increases much more significantly with the increase in the second plate thickness than the cooling cycle time.

[0420] The data indicate that the increase in the sample thickness will result in a significant increase in the energy required to heat and cool the sample.

[0421] For rapid heating and cooling, the sample thickness should be minimized as much as possible. The preferred thickness of the sample is 30 μm or less. Another preferred thickness of the sample is 10 μm or less. Another preferred thickness of the sample is 5 μm or less.

[0422] Experiment 8

[0423] Effect of LED Power on Heating and Cooling Times

[0424] In another experiment, the effect of the LED power on the heating and cooling times was studied. The exemplary RHC card has a first plate of a 50-μm-thick PMMA plate, a second plate of 25-μm-thick PET, and the HC layer is on the outer surface of the second plate. The first plate, the second plate, and the Au / black paint HC layer have the same area of 20 mm × 20 mm. The first plate has a periodic array of spacers on its inner surface with a height of 30 μm, a cross-sectional size of 30 μm × 40 μm, and a spacer pitch of 80 μm. The HC layer contains a 500-nm-thick Au (gold) film and a black paint layer. One surface of the gold film is in contact with the outer surface of the second plate, while the other surface is coated with black paint. The black paint is a commercial product of a film composed of a mixture of black carbon nanoparticles and a polymer. The average thickness of the black paint is -9 μm (-2-μm thickness variation).

[0425] The heating power provided by a blue (450 nm peak wavelength) LED is projected onto a -5 mm × 5 mm heating area of the H / C layer, and the power varies from 100 mW to 500 mW. The sample is a 5 μL liquid temperature-sensitive dye LDS698 at a concentration of 2 mg / mL in 60% water and 40% DMSO. The temperature-sensitive dye allows us to optically measure the sample temperature. The 5 μL sample on the RHC card is much larger than the heating area, with a thickness of 30 μm and an area of -167 mm 2 of area.

[0426] The experimental data shown in Fig. 27A illustrate the relationship between heating time and heating source power, showing the experimental data of the time required to heat from 65 °C to 93 °C with heating LED power intensities from 100 mW to 500 mW on the RHC card.

[0427] The experimental data shown in Fig. 27B illustrate the relationship between cooling time and heating source power, showing the time required to cool from 93 °C to 65 °C. The heating / cooling time results are also shown in Table 1.

[0428] The experimental data indicate that for a given sample holder (i.e., the RHC card), as the LED power increases from 100 mW to 500 mW, the thermal cycle time decreases from 14 seconds to 0.4 seconds, while the cooling cycle time remains almost constant.

[0429] The experimental data indicate that for low heating power (i.e., low heating power density), it will take longer to deliver a fixed amount of energy, and the longer time will increase the energy loss in heat conduction and radiation, and thus increase the wasted energy. For cooling, since the amount of thermal energy stored in a fixed volume of sample at a given temperature is fixed, regardless of how long it takes to reach that temperature, the cooling time is almost independent of the heating cycle time.

[0430] The experiments show that to reduce the total thermal cycle time, the heating power should be increased.

[0431] Table 1. Effect of LED power on RHC heating and cooling times

[0432]

[0433] Experiment 9

[0434] Effect of H / C layer material on heating and cooling times

[0435] In another experiment, the effect of the H / C layer material on the heating and cooling times was studied. An exemplary RHC card has a first plate of a 50-μm thick PMMA sheet, a second plate of a 25-μm thick PET film, a periodic array of spacers that adjust a water-like sample to a 30-μm thickness, and an HC layer on the outer surface of the second plate. The HC layer has different materials for each different RHC card. The first plate, the second plate, and the HC layer have the same area of 20 mm × 20 mm. The LED heating power projected onto a -5 mm × 5 mm heating zone of the H / C layer is 300 mW. A 5-μL water-like sample on the RHC card is much larger than the heating zone area and has a thickness of 30 μm and an area of -167 mm 2 . The thermal cycle is between 65 °C and 93 °C.

[0436] The experimental data in FIGS. 28A and 28B show that for the three different HC layer materials tested, for the sample holder with an HC layer of Au (500 nm thick) plus 9-μm black paint, the heating period and the cooling cycle time are 0.75 s and 0.75 s, respectively, for the sample holder with only an Au (500 nm thick) HC layer, the heating period and the cooling cycle time are 1 s and 1.1 s, respectively, and for the sample holder with an Al (500 nm thick) plus 9-μm black paint HC layer, the heating period and the cooling cycle time are 1.75 s and 1 s, respectively.

[0437] This experiment demonstrates the importance of good lateral thermal conductivity in radiative cooling. Compared with gold with black paint, aluminum plus black paint has almost the same sample light absorption (and thus radiation), but much poorer lateral thermal conductivity, which results in a much smaller effective radiative area because the heat cannot spread laterally as much as possible.

[0438] The experiment shows that a preferred embodiment of the material for the HC layer is a thin gold film plus black paint.

[0439] Experiment 10

[0440] Demonstration of a 0.73-s thermal cycle time (0.23-s heating time and 0.5-s cooling time)

[0441] In another experiment, for a thermal cycle between 65 °C and 93 °C, an RHC card (Card B) was experimentally shown to have a 0.73-s thermal cycle time (0.23-s heating time and 0.5-s cooling time), and another RHC card (Card A) was experimentally shown to have a 0.9-s thermal cycle time (0.3-s heating time and 0.6-s cooling time).

[0442] Figure 29A shows a sample holder with two plates, each plate being a high-density polyethylene (HDPE) film having a thickness of about 10 μm, a width of about 20 mm, and a length of about 20 mm. The spacers that control the sample thickness are soda lime balls having a diameter of about 24 μm at a concentration of about 60 mg / mL. The spherical spacers are mixed with the sample.

[0443] Figure 29B shows a sample holder with a first plate of polymethyl methacrylate (PMMA) film having a thickness of 25 μm and a second plate of high-density polyethylene (HDPE) film having a thickness of 10 μm. The two plates have the same area of 20 mm × 20 mm. The first plate has a periodic array of spacers on its inner surface with a height of 10 μm, a size of 30 μm × 40 μm, and a spacer pitch of 80 μm.

[0444] The two sample holder embodiments shown in Figures 29A and 29B have an H / C layer on the entire outer surface of the second plate. The H / C layer includes an Au film having a thickness of 500 nm, one surface of which is in contact with the outer surface of the second plate, and the other surface is coated with a black coating. The black coating is a commercial product of a film composed of a mixture of black carbon nanoparticles and a polymer, and the coated film has an average thickness of 9 μm and a thickness variation of 2 μm.

[0445] The sample is a liquid temperature-sensitive dye LDS698 at a concentration of 2 mg / mL in 60% water and 40% DMSO. The volume of the sample is 5 μL for the sample holder in Figure 29A and 3 μL for the sample holder in Figure 29B.

[0446] The heat source is a blue light-emitting diode (LED) with a central wavelength of 450 nm, which projects 500 mW of energy onto the black coating layer of the H / C layer, forming a heating zone with an area of -5 mm × 5 mm at the center of the second plate.

[0447] The experimental data (Table 2) show that for thermal cycling from 65 °C to 93 °C, the sample holder of Figure 29A (has a heating cycle time of 0.3 s and a cooling time of 0.60 s, so the total thermal cycle time is 0.90 seconds; the sample holder of Figure 29B has a heating cycle time of 0.23 s and a cooling time of 0.50 s, so the total thermal cycle time is 0.73 seconds.

[0448] Table 2.1 RHC parameters

[0449]

[0450] Table 2.2 Heating / Cooling Performance

[0451]

[0452] Experiment 11

[0453] Effect of Using a Sample Holder and a Sample Adapter on Heating and Cooling Times

[0454] In another experiment, the effect of using a sample holder support and a sample adapter on heating and cooling times was studied.

[0455] In the experiment, an RHC card was placed on a mechanical sample card support (referred to as the "card support"), and then the card support was slid into the adapter. The thermal cycle time was measured for the following cases: (a) only the RHC card, (b) the RHC card on the card support, and (c) the card support and the RHC card on the card support slid into the adapter.

[0456] In some embodiments, the sample holder (shown in FIGS. 30A and 30B) includes a first plate that is a poly(methyl methacrylate) (PMMA) film having a thickness of 10 μm to 50 μm, a width of 22 mm, and a length of 27 mm. The first plate has a periodic array of spacers on its inner surface, the spacers having a height of 30 μm, a cross-sectional size of 30 μm × 40 μm, and a spacer pitch of 80 μm.

[0457] In some embodiments, the second plate is a polyethylene terephthalate (PET) film or a high-density polyethylene film having a thickness of 10 μm to 50 μm, a width of 20 mm, and a length of 27 mm. The heating / cooling layer covers the entire outer surface of the second plate. The heating / cooling layer includes an Au film having a thickness of 100 nm to 500 nm, one surface of the Au film being in contact with the outer surface of the second plate and the other surface being coated with a black coating. The average thickness of the black coating is 9 μm.

[0458] According to some embodiments, the card support (shown in FIGS. 30A and 30B) includes a 1-mm-thick PMMA plate having a width of 24 mm, a length of 32 mm, and a square hole of 15 mm × 15 mm in the center. The RHC card and the card support are bonded together using an adhesive that is 10 to 15 μm thick and is located between the black coating of the RHC card and the surface of the card support, as shown in FIG. 30B.

[0459] According to some embodiments, the card adapter includes an assembly of two U-shaped frames into which the sample card can be slid in or out. One of the U-shaped frames is made of plastic and the other is made of aluminum, where the two U-shaped frames are assembled parallel to each other with a gap therebetween, and the gap is a slot for the sample card to slide. An example of the card adapter is to cut a conventional SD card connector into a U shape (cut from the rear end).

[0460] When testing the thermal cycling time of an RHC card with a 50 nm thick first card and a 25 nm thick PET second card (both having an area of 20 mm × 20 mm), the sample was a 5 μL liquid temperature-sensitive dye LDS698 with a concentration of 2 mg / mL in 60% water and 40% DMSO. A blue 450 nm LED was projected onto a black coating with an area of 5 mm × 5 mm (forming a heating zone) and a power of 300 mW.

[0461] The experimental data shown in Figure 31 indicate that for thermal cycling between 65 °C and 93 °C, (a) for the RHC card only, the heating cycle time was 0.67 s, the cooling cycle was 0.9 s, and the total thermal cycling time was 1.57 s; (b) for the RHC card on the card support, the heating cycle time was 0.77 s, the cooling cycle was 0.87 s, and the total thermal cycling time was 1.64 s; (c) for the RHC card on the card support with the card support slid into the adapter, the heating cycle time was 0.93 s, the cooling cycle was 0.7 s, and the total thermal cycling time was 1.63 s;...

[0462] The experimental data indicate that by cooling the RHC card mainly based on radiative cooling, the RHC card can be supported by the card support and the card support can be inserted into the adapter while increasing the thermal cycling time by less than 4%.

[0463] Experiment 12

[0464] Effect of additional heat sink usage

[0465] In another experiment, the effect of adding an external heat sink on the heating and cooling times was studied. In this experiment, instead of allowing the heat from the RHC card to radiate into the environment, a Peltier cooling device was brought into contact with the edge of the HC layer of the RHC card.

[0466] The RHC card has a first plate of a 50 μm thick PMMA plate, a second plate of a 50 μm thick PET film, a periodic array of spacers that control the sample thickness to 30 μm, and the HC layer is made of bare 0.3 μm thick gold and is located on the outer surface of the second plate. The first plate has an area of 20 mm × 20 mm. The second plate and the HC layer made of gold have the same area of 30 mm × 30 mm.

[0467] The LED heating power projected onto the -5 mm × 5 mm heating zone of the H / C layer was 500 mW. The 5 μL water-like sample on the RHC card was much larger than the heating zone area, having a thickness of 30 μm and an area of -167 mm 2 The thermal cycling was between 65 °C and 93 °C.

[0468] In some setups, the Peltier cooler that provides a 0°C heat sink contacts or is close to the HC layer by overlapping the 3 mm edge with the second plate. In the reference device, there is no Peltier cooler. The sample is a 5 μL liquid temperature-sensitive dye LDS698 at a concentration of 2 mg / mL in 60% water and 40% DMSO.

[0469] The experimental data (Table 3) show that without the Peltier cooler, the liquid in the RHC card takes 0.63 s to heat from 65°C to 93°C, and 1.2 s to cool from 93°C to 65°C, and the Peltier cooler contacts the Au film. The time for the liquid in the RHC card to heat from 65°C to 93°C increases to 0.73 s, while the time to cool from 93°C to 65°C shortens to 0.93 s. Using the Peltier cooler, the total thermal cycle time decreases from 1.83 to 1.66. This is achieved by slightly increasing the heating cycle time and significantly reducing the cooling cycle time.

[0470] Table 3. Effect of using the RHC card with an additional heat sink

[0471]

[0472] Sample card (i.e., RHC card)

[0473] Certain exemplary embodiments of the key components of the sample card (i.e., the RHC card) are given below.

[0474] Sample thickness

[0475] To reduce the thermal mass of the sample and reduce the heat convection loss in the sample, in some embodiments, the average sample thickness at the area heated by the heated / cooled layer is 500 μm or less, 200 μm or less, 100 μm or less, 50 μm or less, 20 μm or less, 10 μm or less, 5 μm or less, 2 μm or less, 1 μm or less, 500 nm or less, 300 nm or less, 100 nm or less, 50 nm or less, or within the range between any two of these values.

[0476] A preferred average sample thickness at the area heated by the heated / cooled layer is 0.1 μm to 0.5 μm, 0.5 μm to 10 μm, 10 μm to 20 μm, 20 μm to 30 μm, 30 μm to 50 μm, 50 μm to 80 μm, 80 μm to 100 μm, or 100 μm to 150 μm.

[0477] Experiment 13

[0478] Example of real-time PCR amplification using the RHC card and system

[0479] The RHC card in this experiment has a first plate of PMMA plate with a thickness of 50 μm, a second plate of PET with a thickness of 25 μm, and the H / C layer is located on the outer surface of the second plate. The gold / black paint HC layer has an area with a diameter of 10 mm. The first plate has a periodic array of spacers on its inner surface. The HC layer contains a thin Au (gold) film and a black paint layer. The gold film has one surface in contact with the outer surface of the second plate and another surface in contact with the black paint. The black paint is a commercial product of a film composed of a mixture of black carbon nanoparticles and polymers. The average thickness of the black paint is -9 μm (-2 μm thickness variation).

[0480] The PCR (real-time PCR) reagent for amplifying the genomic DNA of Staphylococcus aureus with a total volume of 20 μL contains an MSSA forward primer, an MSSA reverse primer, and a Cy5-labeled DNA probe, as well as AptaTaq DNA buffer, AptaTaq polymerase, MgCl 2 , dNTP, bovine serum albumin (BSA), template DNA, and ddH 2 O.

[0481] In the real-time PCR experiment, two positive RHC cards showed a significant increase in the fluorescence signal relative to the number of cycles, especially after 20 cycles of amplification. While the fluorescence signal of a negative RHC card did not show a significant increase relative to the number of cycles. After 40 cycles of amplification in the RHC system, the PCR products in the RHC cards were extracted and subjected to nucleic acid gel electrophoresis to confirm that only the two positive RHC cards had successful amplification bands.

[0482] Sample well

[0483] In some embodiments, one or both plates have sample wells, where the wells regulate the maximum volume of the sample in the well and prevent the sample from flowing into other positions of the plate.

[0484] Plate thickness

[0485] To reduce the thermal mass of the first and second plates and reduce the lateral heat conduction loss in the plates, the thicknesses of the first and second plates are preferably thin.

[0486] In some embodiments, the thickness of the first plate or the second plate is 2 nm or less, 10 nm or less, 100 nm or less, 200 nm or less, 500 nm or less, 1000 nm or less, 2 μm (micrometer) or less, 5 μm or less, 10 μm or less, 20 μm or less, 50 μm or less, 100 μm or less, 150 μm or less, 200 μm or less, 300 μm or less, 500 μm or less, 800 μm or less, 1 mm (millimeter) or less, 2 mm or less, 3 mm or less, 5 mm or less, 10 mm or less, or within the range between any two of these values.

[0487] In some embodiments, the thickness of the first plate or the second plate is 10 nm, 100 nm, 200 nm, 300 nm, 400 nm, 500 nm, 1 μm, 2.5 μm, 5 μm, 10 μm, 25 μm, 50 μm, 100 μm, 200 μm or 500 μm, 1000 μm, or within the range between any two values.

[0488] The first plate and the second plate may have the same thickness or different thicknesses, and may be made of the same material or different materials.

[0489] In some preferred embodiments, the thickness of the first plate or the second plate is between 10 nm and 500 nm, between 500 nm and 1 μm, between 1 μm and 2.5 μm, between 2.5 μm and 5 μm, between 5 μm and 10 μm, between 10 μm and 25 μm, between 25 μm and 50 μm, between 50 μm and 100 μm, between 100 μm and 200 μm or between 200 μm and 500 μm, or between 500 μm and 1000 μm.

[0490] The preferred thickness of the first plate or the second plate is 10 nm or less, 100 nm or less, 200 nm or less, 500 nm or less, 1000 nm or less, 2 μm (micrometer) or less, 5 μm or less, 10 μm or less, 20 μm or less, 50 μm or less, 100 μm or less, 150 μm or less, 200 μm or less, 300 μm or less, 500 μm or less, or within the range between any two of these values.

[0491] In some preferred embodiments, the thickness of the plate with the heating / cooling layer is thinner than that of the other plate without the heater.

[0492] In some preferred embodiments, the thickness of the first plate is 100 nm, 200 nm, 500 nm, 1 μm (micrometer), 2 μm, 5 μm, 10 μm, 25 μm, 50 μm, 100 μm, 125 μm, 150 μm, 175 μm, 200 μm, 250 μm, or within the range between any two of these values; while the thickness of the second plate is 25 μm, 50 μm, 100 μm, 125 μm, 150 μm, 175 μm, 200 μm, 250 μm, 500 μm, 1 mm, 1.5 mm, 2 mm, or within the range between any two of these values.

[0493] In some embodiments, the average thickness of at least one plate is within the range of 1 to 1000 μm, 10 to 900 μm, 20 to 800 μm, 25 to 700 μm, 25 to 800 μm, 25 to 600 μm, 25 to 500 μm, 25 to 400 μm, 25 to 300 μm, 25 to 200 μm, 30 to 200 μm, 35 to 200 μm, 40 to 200 μm, 45 to 200 μm, or 50 to 200 μm.

[0494] In some embodiments, the average thickness of at least one plate is within the range of 50 to 75 μm, 75 to 100 μm, 100 to 125 μm, 125 to 150 μm, 150 to 175 μm or 175 to 200 μm.

[0495] In some embodiments, the average thickness of at least one plate is about 50 μm, about 75 μm, about 100 μm, about 125 μm, about 150 μm, about 175 μm or about 200 μm.

[0496] Plate area. In some embodiments, the lateral area of the first plate and / or the second plate is 1 mm 2 (square millimeter) or less, 10 mm 2 or less, 25 mm 2 or less, 50 mm 2 or less, 75 mm 2 or less, 1 cm 2 (square centimeter) or less, 2 cm 2 or less, 3 cm 2 or less, 3 cm 2 or less, 5 cm 2 or less, 10 cm 2 or less, 20 cm 2 or less, 30 cm 2 or less, 50 cm 2 or less, 100 cm 2 or less, 500 cm 2 or less, 1000 cm2 or less, 5000 cm 2 or less, 10,000 cm 2 or less, or within the range between any two of these values.

[0497] In a preferred embodiment, the lateral area of the first plate and / or the second plate is in the range of 1 mm 2 (square millimeter) to 10 mm 2 , 10 mm 2 to 50 mm 2 , 50 mm 2 to 100 mm 2 , 1 cm 2 to 5 cm 2 , 5 cm 2 to 20 cm 2 , 20 cm 2 to 50 cm 2 , 50 cm 2 to 100 cm 2 , 100 cm 2 to 500 cm 2 , 500 cm 2 to 1000 cm 2 or 1000 cm 2 to 10,000 cm 2 within the range.

[0498] In some embodiments, the first plate and the second plate have the same lateral dimensions. In some embodiments, one of the plates has an area that is 10% or less, 30% or less, 50% or less, 80% or less, 90% or less, 95% or less, 99% or less different from the other plate, or within the range between any two values (taking the largest plate as the basis for calculating the different percentages).

[0499] In some embodiments, the first plate and / or the second plate have a width or length of 5 mm, 10 mm, 20 mm, 25 mm, 30 mm, 40 mm, 50 mm, 75 mm, 100 mm, or within the range between any two of these values.

[0500] In a preferred embodiment, the first plate and / or the second plate have a width or length within the range of 5 mm to 10 mm, 20 mm to 30 mm, 30 mm to 50 mm, 50 mm to 75 mm, or 75 mm to 100 mm.

[0501] In a preferred embodiment, the width or length of the plate is in the range of 5 mm to 50 mm. In another preferred embodiment, the plate has a width within the range of 5 mm to 50 mm and a length within the range of 6 mm to 70 mm.

[0502] Material for the plate

[0503] In some embodiments, the materials for the first plate and the second plate include but are not limited to polymers (such as plastics) or amorphous organic materials. The polymer materials include but are not limited to acrylate polymers, vinyl polymers, olefin polymers, cellulose polymers, non-cellulose polymers, polyester polymers, nylon, cycloolefin copolymers (COC), poly(methyl methacrylate) (PMMA), polycarbonate (PC), cycloolefin polymers (COP), liquid crystal polymers (LCP), polyamides (PA), polyethylene (PE), polyimides (PI), polypropylene (PP), polyphenylene ethers (PPE), polystyrene (PS), polyoxymethylene (POM), polyetheretherketone (PEEK), polyethersulfone (PES), polyethylene terephthalate (PET), polytetrafluoroethylene (PTFE), polyvinyl chloride (PVC), polyvinylidene fluoride (PVDF), polybutylene terephthalate (PBT), fluorinated ethylene propylene (FEP), perfluoroalkoxy alkane (PFA), polydimethylsiloxane (PDMS), rubber, or any combination thereof.

[0504] In some embodiments, the materials for the first plate and the second plate include but are not limited to inorganic materials, which include dielectric materials such as silicon oxide, porcelain, enamel (ceramics), mica, glass, and oxides of various metals.

[0505] In some embodiments, the materials for the first plate and the second plate include but are not limited to inorganic materials, including aluminum oxide, aluminum chloride, cadmium sulfide, gallium nitride, gold chloride, indium arsenide, lithium borohydride, silver bromide, sodium chloride, graphite, carbon nanotubes, carbon fibers, etc.

[0506] In some embodiments, the materials for the first plate and the second plate include but are not limited to metals (such as gold, copper, aluminum, etc.) and alloys.

[0507] In some embodiments, the materials for the first plate and the second plate are made of the multi-layer materials and / or mixtures of materials listed above.

[0508] Heating layer and cooling layer

[0509] In certain embodiments, the heating layer (112-1) and the cooling layer (112-2) include high-K materials and / or high-KC ratio materials. The high-K and / or high-KC ratio materials include materials / structures such as but not limited to metal films, semiconductors, semimetals, plasma surfaces, metamaterials (such as nanostructures), black silicon, graphite, carbon nanotubes, silicon interlayers, graphene, superlattices, plasma materials, any materials / structures that can effectively absorb electromagnetic waves and convert the absorbed energy into heat energy, and any combination thereof.

[0510] For a heating layer heated by an optical heat source, the heating layer includes a material layer that significantly absorbs radiant energy from the optical heat source. Significantly absorbing means that the heating / cooling layer absorbs radiant energy from the optical heat source more significantly than the sample and the plate.

[0511] In some embodiments, the thickness of the heating / cooling layer is in the range of 50 nm to 15 μm. In some embodiments, the heating / cooling layer includes a high-K layer having a thickness in the range of 100 nm to 1 μm.

[0512] In some embodiments, the size of the light heating region is about 1 μm, 2 μm, 5 μm, 10 μm, 20 μm, 50 μm, 100 μm, 200 μm, 500 μm, 1 mm, 2 mm, 5 mm, 10 mm, 20 mm, 50 mm, or 100 mm, or in a range between any two values. In various embodiments, the size and shape of the light heating region can vary.

[0513] In some embodiments, the heating / cooling layer includes an array of coupled dot pillar antennas (D2PAs), such as, but not limited to, the D2PA arrays described in U.S. Provisional Patent Application No. 61 / 347,178 filed on May 21, 2010, U.S. Provisional Patent Application 61 / 622,226 filed on April 10, 2012, PCT Application No. PCT / US2011 / 037455 filed on May 20, 2011, PCT Application No. PCT / US2013 / 032347 filed on March 15, 2013, and U.S. Patent Application No. 13 / 699,270 filed on June 13, 2013, the entire disclosures of which are incorporated herein by reference for all purposes.

[0514] In some embodiments, there can be more than one heating / cooling layer. For example, at least two surfaces of any first or second plate have a heating / cooling layer.

[0515] In some embodiments, the heating / cooling layer can be two layers of material: one for heating and one for cooling, and the two layers of material can be on the same surface of either the first or second plate. For the sample, the heating layer can be on the outer surface of the second plate, while the cooling layer is on the outer or inner surface of the first plate. Even if the cooling layer is on the outer surface of the first plate, as long as the first plate has a thin thickness (e.g., 25 μm or less), this should be effective for cooling the sample.

[0516] Spacer

[0517] In some embodiments of the present invention, there is a spacer between two plates. In some embodiments, at least one spacer is in the sample contact area. In some embodiments, the spacer has a uniform height. In some embodiments, the thickness of the sample is the sample of the spacer height. In some embodiments, the spacer is fixed to one of the plates.

[0518] Spacer function. In the present invention, the spacer is configured to have one or any combination of the following functions and properties: the spacer is configured to (1) control the thickness of the sample or the relevant volume of the sample together with the plate (preferably, the thickness control is precise or uniform or both over the relevant area); (2) enable the sample to have a squeeze-adjusted open flow (CROF) on the plate surface; (3) not occupy a large surface area (volume) in a given sample area (volume); (4) reduce or increase the sedimentation effect of particles or analytes in the sample; (5) change and / or control the wettability of the inner surface of the plate; (6) identify the position, dimensional ratio, and / or information related to the plate, or (7) perform any combination of the above.

[0519] Spacer structure and shape. To achieve the desired reduction and control of the sample thickness, in certain embodiments, the spacers are fixed to their respective plates. Generally, the spacer can have any shape as long as the spacer can adjust the sample thickness during the CROF process, but certain shapes are preferred to achieve certain functions, such as better uniformity, smaller pressing overshoot, etc.

[0520] The spacer is a single spacer or multiple spacers (e.g., an array). Some embodiments of multiple spacers are arrays of spacers (e.g., columns), where the spacer spacing is periodic or aperiodic, or periodic or aperiodic in certain regions of the plate, or has different distances in different regions of the plate.

[0521] There are two types of spacers: open spacers and closed spacers. An open spacer is a spacer that allows the sample to flow through the spacer (i.e., the sample flows around the spacer and through the spacer). For example, a column as a spacer), while a closed spacer is a spacer that blocks the flow of the sample (i.e., the sample cannot flow beyond the spacer, e.g., a ring-shaped spacer, and the sample is within the ring). Both types of spacers use their height to adjust the final sample thickness in the closed configuration.

[0522] In some embodiments, the spacer is only an open spacer. In some embodiments, the spacer is only a closed spacer. In some embodiments, the spacer is a combination of an open spacer and a closed spacer.

[0523] The term "column spacer" refers to a spacer having a columnar shape, and the columnar shape refers to an object having a height and a lateral shape that allows a sample to flow around it during extrusion open flow. In some embodiments, the spacer has a flat top (e.g., a column having a flat top to contact the plate).

[0524] In some embodiments, the lateral shape of the column spacer is a shape selected from the group consisting of: (i) circular, oval, rectangular, triangular, polygonal, annular, star-shaped, letter-shaped (e.g., L-shaped, C-shaped, letters from A to Z), digit-shaped (e.g., shapes similar to 0, 1, 2, 3, 4, … to 9); (ii) shapes in group (i) having at least one rounded corner; (iii) shapes in group (i) having a zigzag or rough edge; and (iv) any superimposition of (i), (ii), and (iii). For multiple spacers, different spacers can have different lateral shapes and dimensions and different distances from adjacent spacers.

[0525] In some embodiments, the spacer can be and / or can include posts, columns, beads, balls, and / or other suitable geometric shapes. The lateral shape and dimensions of the spacer (i.e., transverse to the respective plate surface) can be any shape and dimensions except for the following limitations in some embodiments: (i) the spacer geometry does not cause significant errors in measuring the sample thickness and volume; or (ii) the spacer geometry does not prevent the sample from flowing out between the plates (i.e., it is not a closed form). However, in some embodiments, some spacers need to be closed spacers to restrict sample flow.

[0526] In some embodiments, the shape of the spacer has rounded corners. For example, a rectangular spacer has one, several, or all rounded corners (such as circular instead of 90-degree angles). Rounded corners generally make the manufacture of the spacer easier and cause less damage to biological materials in some cases.

[0527] The sidewalls of the column can be straight, curved, inclined, or of different shapes in different parts of the sidewalls. In some embodiments, the spacer is a column having various lateral shapes, sidewalls, and a ratio of column height to column lateral area.

[0528] In a preferred embodiment, the spacer has the shape of a column for allowing open flow.

[0529] Spacer material. In the present invention, the spacer is generally made of any material that can be used with two plates to adjust the thickness of the relevant volume of the sample. In some embodiments, the material of the spacer is different from the material of the plate. In some embodiments, the material used for the space is at least the same as a part of the material used for at least one plate.

[0530] The spacer is made of a single material, a composite material, multiple materials, multi-layer materials, an alloy, or a combination thereof. Each material used for the spacer is an inorganic material, an organic material, or a mixture, and examples of the materials are given in the paragraphs of Mat-1 and Mat-2. In a preferred embodiment, the spacer is made of the same material as the plate used in the CROF.

[0531] The spacer has mechanical strength and flexibility. In some embodiments, the mechanical strength of these spacers is strong enough such that during the extrusion process of these plates and in the closed configuration, the height of these spacers is the same as or significantly the same as the height of these plates in the open configuration. In some embodiments, the difference between the open configuration and the closed configuration of these spacers can be characterized and predetermined.

[0532] The material used for the spacer is rigid, flexible, or any adaptation between the two. Rigidity is relative to a given pressure used to put the plate in the closed configuration: if the space does not deform more than 1% in height under pressure, the spacer material is considered rigid, otherwise it is flexible. When the spacer is made of a flexible material, the final sample thickness in the closed configuration can still be predetermined based on the pressure and mechanical properties of the spacer.

[0533] Spacers inside the sample. To achieve the desired reduction and control of the sample thickness, especially to achieve good sample thickness uniformity, in certain embodiments, the spacers are placed inside the sample or the relevant volume of the sample. In some embodiments, there is one or more spacers in the sample or the relevant volume of the sample, with a suitable spacer spacing. In certain embodiments, there is at least one spacer inside the sample, at least two spacers inside the sample or the relevant volume of the sample, or at least "n" spacers inside the sample or the relevant volume of the sample, where "n" can be determined by the sample thickness uniformity or the sample flow characteristics required during the CROF.

[0534] The spacer height. In some embodiments, all spacers have the same predetermined height. In some embodiments, the spacers have the same predetermined height. In some embodiments, the spacers can be divided into groups or regions, where each group or region has its own spacer height. And in certain embodiments, the predetermined height of the spacer is the average height of the spacer. In some embodiments, the heights of the spacers are substantially the same. In some embodiments, the percentage of the number of spacers has the same height. In some embodiments, on the same plate, the spacer height in one region is different from the spacer height in another region. In some cases, the plate with different spacer heights in different regions has measured advantages.

[0535] The height of the spacer is selected based on the desired final sample thickness and the remaining sample thickness. The spacer height (predetermined spacer height) and / or the sample thickness is 3 nm or less, 10 nm or less, 50 nm or less, 100 nm or less, 200 nm or less, 500 nm or less, 800 nm or less, 1000 nm or less, 1 μm or less, 2 μm or less, 3 μm or less, 5 μm or less, 10 μm or less, 20 μm or less, 30 μm or less, 50 μm or less, 100 μm or less, 150 μm or less, 200 μm or less, 300 μm or less, 500 μm or less, 800 μm or less, 1 mm or less, 2 mm or less, 4 mm or less, or within a range between any two of these values.

[0536] In a preferred embodiment, the spacer height and / or the sample thickness is from 1 nm to 100 nm, in another preferred embodiment from 100 nm to 500 nm, in a separate preferred embodiment from 500 nm to 1000 nm, in another preferred embodiment from 1 μm (i.e., 1000 nm) to 2 μm, in a separate preferred embodiment from 2 μm to 3 μm, in another preferred embodiment from 3 μm to 5 μm, in a separate preferred embodiment from 5 μm to 10 μm, and in another preferred embodiment from 10 μm to 50 μm, and in a separate preferred embodiment from 50 μm to 100 μm.

[0537] In some embodiments, the spacer height and / or the sample thickness (i) is equal to or slightly greater than the minimum size of the analyte, or (ii) is equal to or slightly greater than the maximum size of the analyte. "Slightly greater" means approximately 1% to 5% greater, and can be any value between the two values.

[0538] In some embodiments, the spacer height, the spacing between the plates, and / or the sample thickness is greater than the minimum size of the analyte (e.g., the analyte has an anisotropic shape), but less than the maximum size of the analyte.

[0539] For example, red blood cells have a disc shape with a minimum size of 2 μm (disc thickness) and a maximum size of 11 μm (disc diameter). In an embodiment of the present invention, the spacer is selected such that the inner surface of the plate in the relevant area is spaced by 2 μm (equal to the minimum size) in one embodiment, 2.2 μm in another embodiment, or 3 (50% larger than the minimum size) in another embodiment, but less than the maximum size of red blood cells. Such embodiments have certain advantages in blood cell counting. In one embodiment, for red blood cell counting, by spacing the inner surface by 2 μm or 3 μm and any value between the two values, an undiluted whole blood sample is confined in this space, and on average, each red blood cell (RBC) does not overlap with other red blood cells, thus allowing for visually accurate counting of red blood cells. (Too much overlap between RBCs may lead to serious errors in counting).

[0540] In the present invention, in some embodiments, when the plate is in a closed configuration, the use of the plate and the spacer not only adjusts the thickness of the sample, but also adjusts the orientation and / or surface density of the analyte / entity in the sample. When the plate is in a closed configuration, the thinner the thickness of the sample, the fewer the analyte / entity per surface area (i.e., the lower the surface concentration).

[0541] Spacer lateral dimensions. For open spacers, the lateral dimensions can be characterized by its lateral dimensions (sometimes called width) in two orthogonal directions x and y. The lateral dimensions of the spacer are the same or different in each direction.

[0542] In some embodiments, the ratio of the lateral dimensions in the x and y directions is 1, 1.5, 2, 5, 10, 100, 500, 1000, 10,000, or within the range between any two values. In some embodiments, different ratios are used to adjust the sample flow direction; the larger the ratio, the flow is along one direction (the larger size direction).

[0543] In some embodiments, the different lateral dimensions of the spacer in the x and y directions are used as (a) using the spacer as a scale marker to indicate the orientation of the plate, (b) using the spacer to generate more sample flow in a preferred direction, or both.

[0544] In a preferred embodiment, period, width, and height.

[0545] In some embodiments, all spacers have the same shape and size. In some embodiments, each spacer has different lateral dimensions.

[0546] For a closed spacer, in some embodiments, the internal lateral shape and dimensions are selected based on the total volume of the sample to be enclosed by the closed spacer, where the volume dimensions have been described in this disclosure; and in certain embodiments, the outer shape and dimensions are selected based on the required strength to support the pressure of the liquid against the spacer and the extrusion pressure of the pressing plate.

[0547] The aspect ratio of the height of the columnar spacer to the average lateral dimension. In certain embodiments, the aspect ratio of the height of the columnar spacer to the average lateral dimension is 100,000, 10,000, 1,000, 100, 10, 1, 0.1, 0.01, 0.001, 0.0001, 0.00001 or within the range between any two of these values.

[0548] Spacer height accuracy. The height of the spacer should be precisely controlled. The relative accuracy of the spacer (i.e., the ratio of the deviation to the desired spacer height) is 0.001% or less, 0.01% or less, 0.1% or less; 0.5% or less, 1% or less, 2% or less, 5% or less, 8% or less, 10% or less, 15% or less, 20% or less, 30% or less, 40% or less, 50% or less, 60% or less, 70% or less, 80% or less, 90% or less, 99.9% or less, or within the range between any values.

[0549] Spacer spacing. The spacer can be a single spacer or multiple spacers on the plate or in the sample-related area. In some embodiments, the spacers on the plate are configured and / or arranged in an array, and the array is periodic, non-periodic, or periodic in some positions of the plate and non-periodic in other positions.

[0550] In some embodiments, the periodic array of spacers is arranged as a lattice of squares, rectangles, triangles, hexagons, polygons, or any combination thereof, where the combination means that different positions on the plate have different spacer lattices.

[0551] In some embodiments, the spacer spacing of the spacer array is periodic in at least one direction of the array (i.e., uniform spacer spacing). In some embodiments, the spacer spacing is configured to improve the uniformity between the plate spacings in the closed configuration.

[0552] The distance between adjacent spacers (i.e., spacer pitch) is 1 μm or less, 5 μm or less, 7 μm or less, 10 μm or less, 20 μm or less, 30 μm or less, 40 μm or less, 50 μm or less, 60 μm or less, 70 μm or less, 80 μm or less, 90 μm or less, 100 μm or less, 200 μm or less, 300 μm or less, 400 μm or less, or within a range between any two values.

[0553] In certain embodiments, the spacer pitch is 400 μm or less, 500 μm or less, 1 mm or less, 2 mm or less, 3 mm or less, 5 mm or less, 7 mm or less, 10 mm or less, or within a range between any values. In certain embodiments, the spacer pitch is 10 mm or less, 20 mm or less, 30 mm or less, 50 mm or less, 70 mm or less, 100 mm or less, or within a range between any values.

[0554] The distance between adjacent spacers (i.e., spacer distance) is selected such that for a given property of the plate and the sample, in the closed configuration of the plate, in some embodiments, the sample thickness variation between two adjacent spacers is at most 0.5%, 1%, 5%, 10%, 20%, 30%, 50%, 80% or within a range between any values; or in certain embodiments, at most 80%, 100%, 200%, 400% or within a range between any two of these values.

[0555] Obviously, in order to maintain a given sample thickness variation between two adjacent spacers, when using a more flexible plate, a closer spacer pitch is required.

[0556] In a preferred embodiment, the spacers are a periodic square array, where the spacers are columns with a height of 2 to 4 μm, an average lateral dimension of 5 to 20 μm, and a spacer pitch of 1 μm to 100 μm.

[0557] In a preferred embodiment, the spacers are a periodic square array, where the spacers are columns with a height of 2 to 4 μm, an average lateral dimension of 5 to 20 μm, and a spacer pitch of 100 μm to 250 μm.

[0558] In a preferred embodiment, the spacers are a periodic square array, where the spacers are columns with a height of 4 to 50 μm, an average lateral dimension of 5 to 20 μm, and a spacer pitch of 1 μm to 100 μm.

[0559] In a preferred embodiment, the spacer is a periodic square array, where the spacer is a pillar with a height of 4 to 50 μm, an average lateral dimension of 5 to 20 μm, and a spacer pitch of 100 μm to 250 μm.

[0560] The period of the spacer array is 1 nm to 100 nm in one preferred embodiment, 100 nm to 500 nm in another preferred embodiment, 500 nm to 1000 nm in a separate preferred embodiment, 1 μm (i.e., 1000 nm) to 2 μm in another preferred embodiment, 2 μm to 3 μm in a separate preferred embodiment, 3 μm to 5 μm in another preferred embodiment, 5 μm to 10 μm in a separate preferred embodiment, and 10 μm to 50 μm in another preferred embodiment, 50 μm to 100 μm in a separate preferred embodiment, 100 μm to 175 μm in a separate preferred embodiment, and 175 μm to 300 μm in a separate preferred embodiment.

[0561] Spacer density. The spacers are at a surface density greater than 1 / μm 2 、 greater than 1 / 10 μm 2 、 greater than 1 / 100 μm 2 、 greater than 1 / 500 μm 2 、 greater than 1 / 1000 μm 2 、 greater than 1 / 5000 μm 2 、 greater than 1 / 0.1 mm 2 、 greater than 1 / 1 mm 2 、 greater than 1 / 5 mm 2 、 greater than 1 / 10 mm 2 、 greater than 1 / 100 mm 2 、 greater than 1 / 1000 mm 2 、 greater than 1 / 10000 mm 2 , or a surface density within the range between any two of these values is arranged on each plate.

[0562] These spacers are configured not to occupy a large surface area (volume) in a given sample area (volume);

[0563] Ratio of spacer volume to sample volume. In many embodiments, the ratio of the spacer volume (i.e., the volume of the spacers) to the sample volume (i.e., the volume of the sample), and / or the ratio of the spacer volume within the relevant volume of the sample to the relevant volume of the sample, is controlled to achieve certain advantages. Advantages include, but are not limited to, uniformity of sample thickness control, uniformity of analytes, and sample flow characteristics (i.e., flow rate, flow direction, etc.).

[0564] In some embodiments, the ratio of the spacer volume (r) to the sample volume, and / or the ratio of the volume of the spacer within the relevant volume of the sample to the relevant volume of the sample is less than 100%, at most 99%, at most 70%, at most 50%, at most 30%, at most 10%, at most 5%, at most 3%, at most 1%, at most 0.1%, at most 0.01%, at most 0.001% or within a range between any values.

[0565] Spacers fixed to the plate. The spacer spacing and the orientation of the spacers, which are crucial in the present invention, are preferably maintained during the process of changing the plate from the open configuration to the closed configuration, and / or are preferably predetermined prior to the process from the open configuration to the closed configuration.

[0566] Some embodiments of the present invention are that, before these plates are changed to the closed configuration, these spacers are fixed to one of the plates. The term "the spacer is fixed together with its corresponding plate" means that the spacer is attached to the plate and maintains this attachment at least during the use of the plate. An example of "the spacer is fixed together with its corresponding plate" is that the spacer is integrally made of a piece of material of the plate, and the position of the spacer relative to the plate surface does not change. An example of "the spacer is not fixed together with its corresponding plate" is that the spacer is adhered to the plate by an adhesive, but during the use of the plate, the adhesive cannot hold the spacer at its original position on the plate surface (i.e., the spacer moves away from its original position on the plate surface).

[0567] In some embodiments, at least one spacer is fixed to its corresponding plate. In certain embodiments, two spacers are fixed to their corresponding plates. In certain embodiments, most of the spacers are fixed with their respective plates. In certain embodiments, all spacers are fixed with their respective plates.

[0568] In some embodiments, the spacer is integrally fixed to the plate.

[0569] In some embodiments, the spacer is fixed to its corresponding plate by one or any combination of the following methods and / or configurations: attached to, joined to, fused to, imprinted and etched.

[0570] The term "imprinted" means that the spacer and the plate are integrally fixed by imprinting (i.e., stamping) a piece of material to form the spacer on the plate surface. The material can be a single-layer material or a multi-layer material.

[0571] The term "etched" means that the spacer and the plate are integrally fixed by etching a piece of material to form the spacer on the plate surface. The material can be a single-layer material or a multi-layer material.

[0572] The term "fusion" means that the spacer and the plate are integrally fixed by attaching the spacer and the plate together, the original materials of the spacer and the plate are fused with each other, and there is a clear material boundary between the two materials after fusion.

[0573] The term "bonded to" means that the spacer and the plate are integrally fixed by adhesive bonding of the spacer and the plate.

[0574] The term "attached to" means that the spacer and the plate are connected together.

[0575] In some embodiments, the spacer and the plate are made of the same material. In other embodiments, the spacer and the plate are made of different materials. In another embodiment, the spacer and the plate are formed as one piece. In another embodiment, one end of the spacer is fixed to its corresponding plate, and the end is open to accommodate different configurations of the two plates.

[0576] In another embodiment, each spacer is independently at least one of attached to, bonded to, fused to, imprinted on, and etched into the corresponding plate. The term "independently" means that one spacer is fixed to its corresponding plate by the same or different methods selected from the methods of attaching to, bonding to, fusing to, imprinting on, and etching into the corresponding plate.

[0577] In some embodiments, at least a segment of the distance between two spacers is predetermined ("predetermined spacer pitch" means that this distance is known when the user uses the plate).

[0578] In some embodiments of all the methods and devices described herein, there are additional spacers in addition to the fixed spacers.

[0579] In a preferred embodiment, the spacer is integrally fabricated on the plate by imprinting (e.g., nanoimprinting) a thin plastic film using a mold, and the spacer is made of the same material, and the thickness of the plate is 50 μm to 500 μm.

[0580] In a preferred embodiment, the spacer is integrally fabricated on the plate by imprinting (e.g., nanoimprinting) a thin plastic film using a mold, and the spacer is made of the same material, and the thickness of the plate is 50 μm to 250 μm.

[0581] In a preferred embodiment, the spacer is integrally fabricated on the plate and is made of the same material, and the thickness of the plate is 50 μm to 500 μm.

[0582] In a preferred embodiment, the spacer is integrally formed as a thin plastic film on the plate using a mold, and is made of the same material, and the thickness of the plate is 50 μm to 250 μm.

[0583] In a preferred embodiment, the spacer is integrally fabricated on a plate by using a mold to imprint (e.g., nanoimprint) a thin plastic film, and the spacer is made of the same material, where the plastic film is PMMA (polymethyl methacrylate) of PS (polystyrene).

[0584] In a preferred embodiment, the spacer is integrally fabricated on a plate by using a mold to imprint (e.g., nanoimprint) a thin plastic film, and the spacer is made of the same material, where the plastic film is PMMA (polymethyl methacrylate) of PS (polystyrene), and the thickness of the plate is from 50 μm to 500 μm.

[0585] In a preferred embodiment, the spacer is integrally fabricated on a plate by using a mold to imprint (e.g., nanoimprint) a thin plastic film, and the spacer is made of the same material, where the plastic film is PMMA (polymethyl methacrylate) of PS (polystyrene), and the thickness of the plate is from 50 μm to 250 μm.

[0586] In a preferred embodiment, the spacer is integrally fabricated on a plate by using a mold to imprint (e.g., nanoimprint) a thin plastic film, and is made of the same material, where the plastic film is PMMA (polymethyl methacrylate) of PS (polystyrene), and the spacer has a square or rectangular shape and has the same spacer height.

[0587] In a preferred embodiment, the spacer has a square or rectangular shape (with or without rounded corners).

[0588] In a preferred embodiment, the spacer has a column width (spacer width in each lateral direction) between 1 μm and 200 μm; a column period (i.e., spacer period) of 2 μm to 2000 μm, and a column height (i.e., spacer height) of 1 μm to 100 μm for a square or rectangular column.

[0589] In a preferred embodiment, the spacer made of PMMA or PS has a column width (spacer width in each lateral direction) between 1 μm and 200 μm; a column period (i.e., spacer period) of 2 μm to 2000 μm, and a column height (i.e., spacer height) of 1 μm to 100 μm for a square or rectangular column.

[0590] In a preferred embodiment, the spacer is integrally fabricated on a plate and made of a plastic material, and the spacer has a column width (spacer width in each lateral direction) between 1 μm and 200 μm; a column period (i.e., spacer period) of 2 μm to 2000 μm, and a column height (i.e., spacer height) of 1 μm to 100 μm for a square or rectangular column.

[0591] In a preferred embodiment, the spacer is integrally fabricated on the plate and made of the same material, and the spacer has square or rectangular pillars with a pillar width (spacer width in each lateral direction) between 1 μm and 200 μm; a pillar period (i.e., spacer period) of 2 μm to 2000 μm, and a pillar height (i.e., spacer height) of 1 μm to 10 μm.

[0592] In a preferred embodiment, the spacer is integrally fabricated on the plate and made of the same material selected from PS or PMMA or other plastics, and the spacer has square or rectangular pillars with a pillar width (spacer width in each lateral direction) between 1 μm and 200 μm; a pillar period (i.e., spacer period) of 2 μm to 2000 μm, and a pillar height (i.e., spacer height) of 10 μm to 50 μm.

[0593] Specific sample thickness. In the present invention, it has been observed that a greater plate holding force (i.e., the force that holds two plates together) can be achieved by using a smaller plate spacing (for a given sample area) or a larger sample area (for a given plate spacing) or both.

[0594] In some embodiments, at least one plate is transparent in the region surrounding the relevant area, each plate having an inner surface configured to contact the sample in the closed configuration; in the closed configuration, the inner surfaces of these plates are substantially parallel to each other; except at the locations where the spacers are present, the inner surfaces of the plates are substantially planar; or any combination thereof.

[0595] Final sample thickness and uniformity. In some embodiments, it is determined to be significantly flat relative to the final sample thickness, and depending on the embodiment and application, the ratio to the sample thickness is less than 0.1%, less than 0.5%, less than 1%, less than 2%, less than 5% or less than 10%, or within a range between any two of these values.

[0596] In some embodiments, the flatness relative to the sample thickness is less than 0.01%, 0.1%, less than 0.1%, less than 0.5%, less than 1%, less than 2%, less than 5%, less than 10%, less than 20%, less than 50%, less than 70%, less than 80%, less than 100%, or within a range between any two of these values.

[0597] In some embodiments, substantially flat may mean that the surface flatness variation itself (measured from the average thickness) is less than 0.1%, less than 0.5%, less than 1%, less than 2%, less than 5%, or less than 10%, or within a range between any two of these values. Generally, the flatness relative to the plate thickness may be less than 0.1%, less than 0.5%, less than 1%, less than 2%, less than 5%, less than 10%, less than 20%, less than 50%, or less than 100%, or within a range between any two of these values.

[0598] The height of the spacer is selected by the required adjusted spacing between the plates and / or the adjusted final sample thickness and the remaining sample thickness. The spacer height (predetermined spacer height), the spacing between the plates, and / or the sample thickness is 3 nm or less, 10 nm or less, 50 nm or less, 100 nm or less, 200 nm or less, 500 nm or less, 800 nm or less, 1000 nm or less, 1 μm or less, 2 μm or less, 3 μm or less, 5 μm or less, 10 μm or less, 20 μm or less, 30 μm or less, 50 μm or less, 100 μm or less, 150 μm or less, 200 μm or less, 300 μm or less, 500 μm or less, 800 μm or less, 1 mm or less, 2 mm or less, 4 mm or less, or within a range between any two of these values.

[0599] The spacer height, the spacing between the plates, and / or the sample thickness is 1 nm to 100 nm in one preferred embodiment, 100 nm to 500 nm in another preferred embodiment, 500 nm to 1000 nm in a separate preferred embodiment, 1 μm (i.e., 1000 nm) to 2 μm in another preferred embodiment, 2 μm to 3 μm in a separate preferred embodiment, 3 μm to 5 μm in another preferred embodiment, 5 μm to 10 μm in a separate preferred embodiment, and 10 μm to 50 μm in another preferred embodiment, 50 μm to 100 μm in a separate preferred embodiment.

[0600] In some embodiments, the spacer can be spherical beads and randomly distributed in the sample.

[0601] In some embodiments, the QMAX device is fully transparent or partially transparent to reduce the heat absorption of the card itself, where the transparency means higher than 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or within a range between any two of these values.

[0602] In some embodiments, the QMAX device is partially reflective to reduce the heat absorption of the card itself. The reflectivity of the surface is higher than 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or within the range between any two values.

[0603] In some embodiments, the QMAX device and the fixture are coated with a thermal insulation layer to reduce the heat absorption of the card itself. The thermal insulation layer comprises a material including the above-mentioned low thermal conductivity material.

[0604] In some embodiments, the fixture covers and seals all QMAX cards in the closed configuration.

[0605] In some embodiments, the fixture only covers and seals the perimeter of the QMAX card in the closed configuration.

[0606] In some embodiments, the fixture only covers and seals the perimeter of the QMAX card in the closed configuration, without covering and sealing the heating and cooling zone area.

[0607] In some embodiments, the fixture covers some surfaces of the QMAX card in the closed configuration.

[0608] In some embodiments, the fixture has a transparent window to allow light to enter and exit the inside of the QMAX card.

[0609] In some embodiments, the fixture is completely transparent to allow light to enter and exit the inside of the QMAX card.

[0610] The transparency of the fixture refers to being higher than 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or within the range between any two values.

[0611] In some embodiments, there is air or liquid between the fixture in the closed configuration and the QMAX device. In certain embodiments, the liquid includes but is not limited to water, ethane, methane, oil, benzene, hexane, heptane, silicone oil, polychlorinated biphenyls, liquid air, liquid oxygen, liquid nitrogen, etc.

[0612] In some embodiments, after closing the fixture, the pressure applied by the fixture on the surface of the QMAX card is 0.01 kg / cm 2 、0.1 kg / cm 2 、0.5 kg / cm 2 、1 kg / cm 2 、2 kg / cm 2 、kg / cm 2 、5 kg / cm 2 、10 kg / cm 2 、20 kg / cm 2, 30 kg / cm 2 , 40 kg / cm 2 , 50 kg / cm 2 , 60 kg / cm 2 , 100 kg / cm 2 , 150 kg / cm 2 , 200 kg / cm 2 , 500 kg / cm 2 , or within a range between any two values; and 0.1 kg / cm 2 to 0.5 kg / cm 2 , 0.5 kg / cm 2 to 1 kg / cm 2 , 1 kg / cm 2 to 5 kg / cm 2 , 5 kg / cm 2 to 10 kg / cm 2 (pressure) preferred range.

[0613] In some embodiments, after closing the clamp, the pressure applied by the clamp on the QMAX card surface is at least 0.01 kg / cm 2 , 0.1 kg / cm 2 , 0.5 kg / cm 2 , 1 kg / cm 2 , 2 kg / cm 2 , kg / cm 2 , 5 kg / cm 2 , 10 kg / cm 2 , 20 kg / cm 2 , 30 kg / cm 2 , 40 kg / cm 2 , 50 kg / cm 2 , 60 kg / cm 2 , 100 kg / cm 2 , 150 kg / cm 2 , 200 kg / cm 2 or 500 kg / cm 2 ,

[0614] As shown in the cross-sectional views of the device in FIGS. 2A and 2B, the heating / cooling layer 112 spans the sample contact area. However, it should be noted that the lateral area of the heating / cooling layer can also occupy only a part of the sample contact area, and the percentage is about 1% or more, 5% or more, 10% or more, 20% or more, 50% or more, 80% or more, 90% or more, 95% or more, 99% or more, 85% or less, 75% or less, 55% or less, 40% or less, 25% or less, 8% or less, 2.5% or less. In some embodiments, in order to facilitate the temperature change of the sample, in some embodiments, the lateral area of the heating / cooling layer is configured such that the sample 90 receives thermal radiation from the heating / cooling layer 112, and the thermal radiation is substantially uniformly distributed across the lateral dimension of the sample 90 on the sample contact area.

[0615] In some embodiments, the radiation absorption area is 10%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100% of the total plate area, or within a range between any two values.

[0616] In some embodiments, the thickness of the heating / cooling layer 112 is 10 nm or greater, 20 nm or greater, 50 nm or greater, 100 nm or greater, 200 nm or greater, 500 nm or greater, 1 μm or greater, 2 μm or greater, 5 μm or greater, 10 μm or greater, 20 μm or greater, 50 μm or greater, 100 μm or greater, 75 μm or less, 40 μm or less, 15 μm or less, 7.5 μm or less, 4 μm or less, 1.5 μm or less, 750 μm or less, 400 nm or less, 150 nm or less, 75 nm or less, 40 nm or less, or 15 nm or less, or within a range between any two values. In certain embodiments, the thickness of the heating / cooling layer 112 is 100 nm or less.

[0617] In some embodiments, the area of the sample layer and the heating / cooling layer 112 is substantially greater than the uniform thickness. Here, the term "substantially greater" means that the overall diameter or diagonal distance of the sample layer and / or the heating layer / cooling layer is at least 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 55 times, 60 times, 65 times, 70 times, 75 times, 80 times, 85 times, 90 times, 95 times, 100 times, 150 times, 200 times, 250 times, 300 times, 350 times, 400 times, 450 times, 500 times, 550 times, 600 times, 650 times, 700 times, 750 times, 800 times, 850 times, 900 times, 950 times, 1000 times, 1500 times, 2000 times, 2500 times, 3000 times, 3500 times, 4000 times, 4500 times, or 5000 times, or within a range between any two values.

[0618] Figures 11A and 11B illustrate exemplary embodiments of the first plate and the heating / cooling layer of the present invention. Figure 11A is a top view, while Figure 11B is a cross-sectional view. Figures 12A and 12B illustrate cross-sectional views of two exemplary embodiments of the present invention, demonstrating the first plate, the second plate, and the heating / cooling layer. As a whole, the first plate and the second plate, and optionally the heating / cooling layer, can be regarded as a sample holder, which relates not only to the embodiments shown and / or described herein, but also to other embodiments capable of pressing at least a part of a liquid sample into a layer with a uniform thickness.

[0619] As shown in Figures 11A and 11B, in some embodiments, the heating / cooling layer is in contact with the first plate. However, it should be noted that in some embodiments, the heating / cooling layer can be in contact with the second plate 20. Additionally, in some embodiments, the heating / cooling layer is not in contact with any plate. In some embodiments, there is no separate structure for the heating / cooling layer; the first plate and / or the second plate 20 and / or the sample itself can absorb electromagnetic radiation, such that the temperature of the sample can increase.

[0620] In some embodiments, the area of the heating / cooling layer is less than 1000 mm 2 、900 mm 2 、800 mm 2 、700 mm 2 、600 mm 2 、500 mm 2 、400 mm 2 、300 mm 2 、200 mm 2 、100 mm 2 、90 mm 2 、80 mm 2 、75 mm 2 、70 mm2 , 60 mm 2 , 50 mm 2 , 40 mm 2 , 30 mm 2 , 25 mm 2 , 20 mm 2 , 10 mm 2 , 5 mm 2 , 2 mm 2 , 1 mm 2 , 0.5 mm 2 , 0.2 mm 2 , 0.1 mm 2 or 0.01 mm 2 , or within a range between any two values. In some embodiments, the heating / cooling layer has an area that is substantially less than the area of the first plate (and / or the second plate). For example, in certain embodiments, the area of the heating / cooling layer only occupies a percentage of the area of the first plate (or the second plate; or the sample contact area of the first plate or the second plate) that is about 1% or greater, 5% or greater, 10% or greater, 20% or greater, 50% or greater, 80% or greater, 90% or greater, 95% or greater, 99% or greater, 85% or less, 75% or less, 55% or less, 40% or less, 25% or less, 8% or less, 2.5% or less.

[0621] In some embodiments, the heating / cooling layer has a substantially uniform thickness. In some embodiments, the thickness of the heating / cooling layer is less than 10 nm, 20 nm, 50 nm, 100 nm, 200 nm, 500 nm, 1 μm, 2 μm, 5 μm, 10 μm, 20 μm, 50 μm, 100 μm, 200 μm, 300 μm, 400 m, 500 m, 600 m, 700 m, 800 m, 900 m, 1 mm, 1.5 mm, 2 mm, 2.5 mm, 3 mm, 3.5 mm, 4 mm, 4.5 mm, 5 mm or 10 mm, or within a range between any two values.

[0622] The heating / cooling layer can adopt any shape. For example, when viewed from a top view, the heating / cooling layer can be square, circular, oval, triangular, rectangular, parallelogram, trapezoidal, pentagonal, hexagonal, octagonal, polygonal or various other shapes.

[0623] In some embodiments, the thickness of the first plate or the second plate is 2 nm or less, 10 nm or less, 100 nm or less, 200 nm or less, 500 nm or less, 1000 nm or less, 2 μm (micrometer) or less, 5 μm or less, 10 μm or less, 20 μm or less, 50 μm or less, 100 μm or less, 150 μm or less, 200 μm or less, 300 μm or less, 500 μm or less, 800 μm or less, 1 mm (millimeter) or less, 2 mm or less, 3 mm or less, 5 mm or less, 10 mm or less, 20 mm or less, 50 mm or less, 100 mm or less, 500 mm or less, or within a range between any two of these values.

[0624] In some embodiments, the lateral area of the first plate and the second plate is 1 mm 2 (square millimeter) or less, 10 mm 2 or less, 25 mm 2 or less, 50 mm 2 or less, 75 mm 2 or less, 1 cm 2 (square centimeter) or less, 2 cm 2 or less, 3 cm 2 or less, 4 cm 2 or less, 5 cm 2 or less, 10 cm 2 or less, 100 cm 2 or less, 500 cm 2 or less, 1000 cm 2 or less, 5000 cm 2 or less, 10,000 cm 2 or less, 10,000 cm 2 or less, or within a range between any two of these values.

[0625] In certain embodiments, the spacer pitch (ISD) of the spacer divided by the fourth power of the thickness (h) and Young's modulus (E) of the plate (ISD 4 / (hE)) is 5 × 10 6 μm 3 / GPa or less;

[0626] In certain embodiments, the product of the pillar contact filling factor and the Young's modulus of the spacer is 2 MPa or greater, where the pillar contact filling factor is the ratio of the area of the plate being pillar contacted to the entire plate area (in the pillar region).

[0627] In some embodiments, the spacer has a predetermined substantially uniform height and a predetermined constant spacer pitch that is at least about 2 times greater than the size of the analyte, up to 200 μm, and wherein at least one spacer is located within the sample contact area.

[0628] In some embodiments, the plate (the first plate, the second plate, or both plates) having the heating / cooling layer is thin such that the temperature of the sample can be changed rapidly. For example, in certain embodiments, the thickness of the plate in contact with the heating / cooling layer is equal to or less than 500 μm, 200 μm, 100 μm, 50 μm, 25 μm, 10 μm, 5 μm, 2.5 μm, 1 μm, 500 nm, 400 nm, 300 nm, 200 nm, or 100 nm, or in a range between any two values. In some embodiments, if only one plate is in contact with the heating / cooling layer, the plate in contact with the heating / cooling layer is substantially thinner than the plate not in contact with the heating / cooling layer. For example, in some embodiments, the thickness of the plate in contact with the heating / cooling layer is less than 1 / 1,000,000, 1 / 500,000, 1 / 100,000, 1 / 50,000, 1 / 10,000, 1 / 5,000, 1 / 1,000, 1 / 500, 1 / 100, 1 / 50, 1 / 10, 1 / 5, or 1 / 2 of the thickness of the plate in contact with the heating / cooling layer, or in a range between any two values.

[0629] In some embodiments, the sample layer is thin such that the temperature of the sample layer can be changed rapidly. In certain embodiments, the thickness of the sample layer is equal to or less than 100 μm, 50 μm, 25 μm, 10 μm, 5 μm, 2.5 μm, 1 μm, 500 nm, 400 nm, 300 nm, 200 nm, or 100 nm, or in a range between any two values.

[0630] In various embodiments, the positioning of the heating / cooling layer can also vary. In some embodiments, as shown in FIGS. 12A or 12B, the heating / cooling layer is located at the inner surface of the first plate. Here, the inner surface is defined as the surface that contacts the sample when the sample is squeezed into a layer. The other surface is the outer surface. In some embodiments, the heating / cooling layer is at the inner surface of the first plate. In some embodiments, the heating / cooling layer is at the inner surface of the second plate. In some embodiments, the heating / cooling layer is at the outer surface of the first plate. In some embodiments, the heating / cooling layer is inside one or both plates. In some embodiments, the heating / cooling layer is at the outer surface of the second plate. In some embodiments, there are at least two heating / cooling layers at the inner surface and / or the outer surface of the first plate and / or the second plate.

[0631] As shown and described herein, in some embodiments, the sample holder is configured to squeeze a fluid sample into a thin layer, thereby reducing the thermal mass of the sample. However, if the thermal mass is reduced, a small amount of energy can rapidly change the temperature of the sample. In addition, by restricting the sample thickness, heat conduction is also restricted.

[0632] In some embodiments, there are sample contact regions on the respective surfaces of the first plate 10 and the second plate 20. The sample contact region can be any part of the surface of the first plate 10 and / or the second plate 20. In some embodiments, the heating / cooling layer at least partially overlaps with the sample contact region. In the overlapping portion, due to being close to each other and having a small thermal mass, the sample is rapidly heated.

[0633] In some embodiments, the sample holder 100 is an extrusion-adjusted open flow (CROF, also known as QMAX) device, such as but not limited to the CROF devices described in the following patent applications: U.S. Provisional Patent Application No. 62 / 202,989 filed on August 10, 2015, U.S. Provisional Patent Application No. 62 / 218,455 filed on September 14, 2015, U.S. Provisional Patent Application No. 62 / 293,188 filed on February 9, 2016, U.S. Provisional Patent Application No. 62 / 305,123 filed on March 8, 2016, U.S. Provisional Patent Application No. 62 / 369,181 filed on July 31, 2016, U.S. Provisional Patent Application No. 62 / 394,753 filed on September 15, 2016, PCT Application (designating the United States) No. PCT / US2016 / 045437 filed on August 10, 2016, PCT Application (designating the United States) No. PCT / US2016 / 051775 filed on September 14, 2016, PCT Application (designating the United States) No. PCT / US2016 / 051794 filed on September 15, 2016, and PCT / US2016 / 054025 filed on September 27, 2016, the entire disclosures of which are incorporated herein by reference for all purposes.

[0634] Edge sealing for reducing sample evaporation

[0635] When the two plates sandwich the sample in a shape with a large lateral / vertical ratio (e.g., 15 mm vs. 30 μm = 500), the evaporation of the sample during thermal cycling is greatly reduced because the sample surface covered by the two plates is 500 times larger. Experiments have found that there is no visible change in the sample volume during 30 temperature cycles (about 60 seconds).

[0636] In another aspect, in some embodiments, there is a sealing element that contacts two plates to form a closed chamber that prevents the outflow of sample vapor. In addition to reducing or eliminating sample evaporation, such a sealing element can also reduce sample contamination. The sealing element can be a band, a plastic seal, an oil seal, or a combination thereof.

[0637] In some embodiments, the sealing element does not reach the sample, but the sealing element contacts two plates to form a closed chamber that prevents the outflow of sample vapor. In some embodiments, the sealing element can be used as a spacer to adjust the thickness of the relevant sample.

[0638] In some embodiments, as shown in FIG. 7, the sample holder 100 includes a sealing element 30 that is configured to seal the spacing 102 between the first plate 10 and the second plate 20 outside the medium contact area in a closed configuration. In certain embodiments, the sealing element 30 encloses the sample 90 within a certain area (e.g., the sample receiving area) such that the entire lateral area of the sample 90 is clearly defined and measurable. In certain embodiments, the sealing element 30 improves the uniformity of the sample 90, particularly the thickness of the sample layer.

[0639] In some embodiments, as shown in FIG. 7, the sealing element 30 includes an adhesive that is coated between the first plate 10 and the second plate 20 in a closed configuration. The adhesive is selected from materials such as, but not limited to: starch, dextrin, gelatin, tar, asphalt, polyisoprene natural rubber, resin, shellac, cellulose and its derivatives, vinyl derivatives, acrylic derivatives, reactive acrylic groups, polychloroprene, styrene-butadiene, styrene-diene-styrene, polyisobutylene, acrylonitrile-butadiene, polyurethane, polysulfide, silicone, aldehyde condensation resin, epoxy resin, amino resin, polyester resin, polyolefin polymer, soluble silicate, phosphate cement, or any other adhesive material, or any combination thereof. In some embodiments, the adhesive is a dry adhesive, a pressure-sensitive adhesive, a contact adhesive, a heat adhesive, or a one-component or multi-component reactive adhesive, or any combination thereof. In some embodiments, the adhesive is a natural adhesive or a synthetic adhesive, or from any other source, or any combination thereof. In some embodiments, the adhesive is self-curing, heat-curing, UV-curing, or cured by any other treatment, or any combination thereof.

[0640] In some embodiments, as shown in FIG. 7, the sealing element 30 includes a closing spacer (hole). For example, when viewed from a top view, the closing spacer has a circular shape (or any other closed shape) and surrounds the sample 90, substantially confining the sample 90 together with the first plate 10 and the second plate 20. In certain embodiments, the closing spacer (hole) also serves as a spacing mechanism 40. In such embodiments, the closing spacer seals the lateral boundaries of the sample 90 and adjusts the thickness of the sample layer.

[0641] In some embodiments, during heating, there is an "anti-evaporation ring" outside the liquid region (e.g., the sample region), which prevents or reduces the escape of the vapor of the liquid from the card.

[0642] In some embodiments, there is a clamp outside the QMAX card to fix the QMAX card in its closed configuration during heating.

[0643] In some embodiments, the two plates are squeezed with an inexact pressure, neither setting the pressure to an exact level nor making it substantially uniform. In certain embodiments, the two plates are directly pressed by hand.

[0644] In some embodiments, the QMAX card / RHC including the plates and the spacer is made of a material with low thermal conductivity to reduce the heat absorption of the card itself.

[0645] In some embodiments, there is a clamp outside the QMAX card to fix the QMAX card in its closed configuration during heating (i.e., the clamp only surrounds the edges of the plates, rather than the center of the plate pair). Wherein the clamp is also made of a material with low thermal conductivity to reduce the heat absorption of the card itself.

[0646] Heating source, additional heat sink, temperature sensor and temperature control

[0647] The heating layer or the heating / cooling layer in the RHC card is configured to be heated by a heating source, wherein the heating source transfers thermal energy to the heating / cooling layer through light, electricity, radio frequency (RF) radiation or a combination thereof.

[0648] Optical heating source. In some embodiments, when the heating layer is optically heated by a heating source, the heating source includes a light source, which includes but is not limited to LEDs (light emitting diodes), lasers, lamps or a combination thereof.

[0649] In order to allow more light from the light source in the optical heating source to reach the heating layer, some embodiments of the heating source use optical lenses, light guides or a combination thereof.

[0650] In some embodiments, the wavelength of the electromagnetic wave is 50 nm, 100 nm, 150 nm, 200 nm, 250 nm, 300 nm, 350 nm, 400 nm, 450 nm, 500 nm, 550 nm, 600 nm, 650 nm, 700 nm, 750 nm, 800 nm, 850 nm, 900 nm, 950 nm, 1 μm, 10 μm, 25 μm, 50 μm, 75 μm, or 100 μm, or within a range between any two values. In some embodiments, the wavelength of the electromagnetic wave is from 100 nm to 300 nm, from 400 nm to 700 nm (visible range), from 700 nm to 1000 nm (IR range), from 1 μm to 10 μm, from 10 μm to 100 μm, or within a range between any two values.

[0651] The NA (numerical aperture) of the lens is 0.001, 0.01, 0.05, 0.1, 0.2, 0.3, 04, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.5, or within a range between any two values.

[0652] In a preferred embodiment, the NA of the lens is from 0.01 to 0.1, from 0.1 to 0.4, from 0.4 to 0.7, from 0.7 to 1.0 or from 1.0 to 1.5.

[0653] FIGS. 14A and 14B respectively show a perspective view and a cross-sectional view of an embodiment of the present invention, in which an optical waveguide is used to guide electromagnetic waves (e.g., LED light) from a heat source (e.g., a light source) to a heating zone or plate.

[0654] In certain embodiments, collimating the light from the light source into the optical waveguide (also referred to as an optical collimator) in the heating zone / plate includes a hollow tube having a reflective wall.

[0655] One embodiment of the optical waveguide includes a hollow dielectric tube having a reflective wall (i.e., its inner wall, outer wall, or both are reflective). The hollow dielectric tube can be made of a material such as glass, plastic, or a combination thereof. The reflective wall can be a thin light-reflective coating on the hollow tube wall. The reflective coating can be a thin metal film, such as gold, aluminum, silver, copper, or any mixture or combination thereof. FIG. 17 shows a perspective view of an embodiment of the optical waveguide that includes a hollow tube and a reflective material coated on the outer wall of the tube. The reflective coating can also be in the inner wall. The reflective wall can also be made of a multi-layer interference material that reflects the relevant light. The optical waveguide can be a block of material having a hollow tube with a reflective wall.

[0656] In some embodiments, the hollow tube has a length in the range of 1 mm to 70 mm, an internal dimension (diameter or width) in the range of 1 mm to 40 mm, and a wall thickness in the range of 0.01 mm to 10 mm.

[0657] In some preferred embodiments, the inner diameter (or average width) of the hollow tube for the light guide tube is in the range of 1 mm to 5 mm, 5 mm to 10 mm, 10 mm to 15 mm, 15 mm to 20 mm, 20 mm to 30 mm, or 30 mm to 50 mm.

[0658] In some preferred embodiments, the wall thickness (or average width) of the hollow tube for the light guide tube is in the range of 0.001 mm to 0.01 mm, 0.01 mm to 0.1 mm, 0.1 mm to 0.5 mm, 0.5 mm to 1 mm, 1 mm to 2 mm, or 2 mm to 50 mm.

[0659] Electric heating source. In some embodiments, when the heating layer or the heating / cooling layer is electrically heated by a heating source, the electric heating source includes a power supply that sends electrical energy to the heating / cooling layer through wires.

[0660] Additional radiator. In some embodiments, heat is removed from the sample and the sample holder to the environment, but in some embodiments, an additional radiator will be used to accelerate heat removal. The additional radiator can be a Peltier cooler, a passive radiator, or both. In some embodiments, a fan will be used to generate air convection that accelerates the cooling of the sample (directly to the sample and the sample holder, directly to the additional radiator, or both).

[0661] Figures 6A and 6B also respectively show a perspective view and a cross-sectional view of some embodiments of a thermal cycling system that includes a sample holder 100 and a thermal control unit 200 in a closed position. The sample holder 100 may include a first plate 10, a second plate 20, and a spacer mechanism (not shown). The thermal control unit 200 may include a heating source 202 and a controller 204.

[0662] As shown in FIG. 6B, the thermal control unit 200 may include a heating source 202 and a controller 204. In some embodiments, the thermal control unit 200 provides energy in the form of electromagnetic waves for changing the temperature of the sample.

[0663] Referring to both FIGS. 6A and 6B, the heating source 202 is configured to project electromagnetic waves 210 onto the heating / cooling layer 112 of the sample holder 100, and the heating / cooling layer 112 is configured to absorb the electromagnetic waves 210 and convert most of the electromagnetic waves 210 into heat to generate thermal radiation that raises the temperature of the portion of the sample 90 close to the heating / cooling layer 112. In other words, the coupling of the heating source 202 and the heating / cooling layer 112 is configured to provide the thermal energy required to promote the temperature change of the sample 90.

[0664] In some embodiments, the radiation from the heating source 202 includes radio waves, microwaves, infrared waves, visible light, ultraviolet waves, X-rays, gamma rays, or thermal radiation, or any combination thereof. In some embodiments, the heating / cooling layer 112 has a preferred optical wavelength range within which the absorption efficiency of the heating / cooling layer 112 is high. In some embodiments, the heating source 202 is configured to project electromagnetic waves in a wavelength range that is within, overlaps with, or encompasses the preferred wavelength range of the heating / cooling layer 112. In other embodiments, to facilitate temperature change, the wavelength is rationally designed to be away from the preferred wavelength of the heating / cooling layer.

[0665] In some embodiments, the heating source 202 includes a laser source that provides laser light in a narrow wavelength range. In other embodiments, the heating source 202 includes one LED among LEDs (light-emitting diodes).

[0666] Temperature sensor. The temperature of the sample can be controlled by delivering pre-calibrated energy to the heating zone / layer with a real-time temperature sensor, by using a real-time temperature sensor, or both.

[0667] The real-time temperature sensor can be a thermometer, a thermocouple, a radiation temperature sensor, a temperature-sensitive dye (which changes light intensity or color or both with temperature), or a combination thereof.

[0668] As shown in FIG. 7, in some embodiments, the thermal control unit 200 includes a thermometer 206. In some embodiments, the thermometer 206 provides a monitoring and / or feedback mechanism to control / monitor / regulate the temperature of the sample 90. For example, in some embodiments, the thermometer 206 is configured to measure the temperature at or near the sample contact area. In certain embodiments, the thermometer 206 is configured to directly measure the temperature of the sample 90. In some embodiments, the thermometer 206 is selected from fiber optic thermometers, infrared thermometers, liquid crystal thermometers, pyrometers, quartz thermometers, silicon bandgap temperature sensors, temperature strips, thermistors, and thermocouples. In certain embodiments, the thermometer 206 is an infrared thermometer.

[0669] In some embodiments, thermometer 206 is configured to send a signal to controller 204. Such a signal contains information related to the temperature of sample 90, enabling controller 204 to make corresponding changes. For example, during a PCR process, for the denaturation step, the target temperature is set to 95°C; after measurement, the thermometer sends a signal to controller 204 indicating that the measured temperature of sample 90 is actually 94.8°C; controller 204 then changes the output of heating source 202, which projects electromagnetic waves or adjusts specific parameters (such as intensity or frequency) of existing electromagnetic waves, so that the temperature of sample 90 increases to 95°C. This measurement-signal adjustment cycle is applied to any step in any reaction / assay.

[0670] Controller. Referring to FIGS. (A) and (B) of FIG. 4, controller 204 is configured to control electromagnetic wave 210 projected by heating source 202 for changing the temperature of the sample. Parameters of electromagnetic wave 210 controlled by controller 204 include but are not limited to: presence, intensity, wavelength, incident angle, and any combination thereof. In some embodiments, the controller is manually operated. For example, it simply controls the turning on and off of the heating source, similar to a manual switch, thereby controlling the presence or absence of the electromagnetic wave projected by the heating source. In other embodiments, the controller includes hardware and software configured to automatically control the electromagnetic wave according to one or more predetermined programs.

[0671] In some embodiments, the predetermined program refers to a schedule where parameters of electromagnetic wave 210 (such as presence, intensity, and / or wavelength) are set to predetermined levels for corresponding predetermined time periods. In other embodiments, the predetermined program refers to a schedule where the temperature of sample 90 is set to a predetermined level for corresponding predetermined time periods, and the time periods for the sample temperature to change from one predetermined level to another are also set respectively. In some embodiments, controller 204 is configured to be programmable, which means that controller 204 includes hardware and software configured to receive and execute the predetermined program of the system, which is provided by the operator of the system.

[0672] FIG. 7 shows a cross-sectional view of an embodiment of the present invention, demonstrating a thermal cycler system and showing additional elements that facilitate temperature change and control. As shown in FIG. 7, the thermal cycler system includes a sample holder 100 and a thermal control unit 200. Sample holder 100 includes a first plate 10, a second plate 20, a spacer mechanism 40, and a sealing element 30; thermal control unit 200 includes a heating source 202, a controller 204, a thermometer 206, and an expander 208.

[0673] Figure 7 shows the sample holder 100 in a closed configuration, where the inner surfaces 11 and 21 of the first plate 10 and the second plate 20 face each other, and the spacing 102 between the two plates is adjusted by the spacer mechanism 40. If the sample 90 has been deposited on one or both plates in the open configuration, when switching to the closed configuration, the first plate 10 and the second plate 20 are pressed by hand or other mechanism, and the sample 90 is thus squeezed into a thin layer by the two plates. In some embodiments, the thickness of the layer is uniform and the same as the spacing 102 between the two plates. In certain embodiments, the spacing 102 (and thus also the thickness of the sample layer) is adjusted by the spacer mechanism 40. In some embodiments, the spacer mechanism includes a closed spacer fixed to one of the plates. In some embodiments, the spacer mechanism 40 includes a plurality of columnar spacers fixed to one or both plates. Here, the term "fixed" means that the spacer is attached to the plate and maintains this attachment at least during the use of the plate.

[0674] In some embodiments, the controller 204 is configured to adjust the temperature of the sample according to a predetermined program to facilitate the determination and / or reaction involving the sample 90. In some embodiments, the determination and / or reaction is PCR. In certain embodiments, the controller 204 is configured to control the presence, intensity, and / or frequency of the electromagnetic wave from the heating source 206.

[0675] Sample signal monitoring

[0676] As shown in FIGS. 11 and 12, a signal sensor can be used to detect the signal of the sample (and the reaction product during the temperature change) in the sample holder.

[0677] In some embodiments, the signal sensor is an optical sensor configured to image the fluid sample. For example, the optical sensor is a photodetector, a camera, or a device capable of capturing an image of the fluid sample. In some embodiments, the optical sensor can be a camera. In some embodiments, the camera is a camera integrated into a mobile device (such as a smartphone or a tablet computer). In some embodiments, the camera is separated from other parts of the system. In some embodiments, one or more light sources are used to excite the sample (and the product from the reaction during the temperature change) to generate a signal.

[0678] In some embodiments, the signal sensor is an electrical sensor configured to detect the electrical signal from the device. In some embodiments, the signal sensor is a mechanical sensor configured to detect the mechanical signal from the device.

[0679] In some embodiments, the signal sensor is configured to monitor the amount of analyte in the sample. In some embodiments, the signal sensor is located outside the chamber and receives the optical signal from the sample through the optical aperture on the chamber.

[0680] Base and system

[0681] In some embodiments, the device further includes a base (adapter) configured to accommodate a sample card, a heating source, a temperature sensor, a part of the overall temperature control (including a smartphone in some embodiments), an additional heat sink (optionally), a fan (optionally), or a combination thereof. In some embodiments, the adapter includes a card slot into which the sample card can be inserted. In some embodiments, after the sample card is fully inserted into the slot or reaches a predetermined position in the slot, it is stabilized and held in place without any movement.

[0682] In some embodiments, the base (adapter) is configured to position the sample card and the sample within the sample card in the field of view of an optical sensor (e.g., a camera) such that the sample can be imaged. In certain embodiments, the camera is part of a mobile device (e.g., a smartphone). In some embodiments, the adapter includes a slider in a slot. In certain embodiments, the sample card can be placed on the slider, and the slider can slide into or out of the slot in the adapter. In some embodiments, the adapter includes a card support. In certain embodiments, the sample card can be placed on the card support, which does not need to be moved prior to imaging.

[0683] In some embodiments, the adapter is configured to be connectable to an optical sensor such that the optical sensor (e.g., a mobile device; e.g., a smartphone) and the sample card are fixed. In certain embodiments, the adapter can include a replaceable connection member (by way of example) that attaches directly to the mobile device. The connection member can slide onto the mobile device and securely attach the adapter to the mobile device, optimally positioning the sample card to be imaged or for the detection and / or measurement of an analyte. In certain embodiments, the connection member is replaceable such that different connection members can be used for different mobile devices.

[0684] In some embodiments, the adapter includes a radiation aperture that allows electromagnetic waves for heating or cooling the sample to pass through. In some embodiments, the adapter includes an optical aperture that allows the sample to be imaged. In some embodiments, the adapter serves as a heating slot for the sample card. Figures 13 and 14 provide other embodiments of the system. Figure 13 shows a cross-sectional view of an exemplary embodiment of the present invention demonstrating a system for rapidly changing the temperature of a sample. Figure 13 shows the detailed elements of a heating source according to one embodiment.

[0685] As shown in FIGS. 13 and 14, in some embodiments, the system includes a sample holder and a heating source. In some embodiments, as described herein, the sample holder includes a first plate, a second plate, and / or a heating / cooling layer. The heating source emits electromagnetic waves that reach the sample and can be converted into heat that raises the temperature of the sample. In some embodiments, the conversion is carried out by the heating / cooling layer. When there is no specific heating / cooling layer, the conversion is carried out by other parts of the sample holder.

[0686] As shown in FIGS. 13 and 14, in some embodiments, the system includes a chamber that encloses the sample holder. In some embodiments, the chamber is an example of the additional radiator in FIG. 1. In some embodiments, the chamber includes an optical aperture that is configured to allow imaging of the sample. In some embodiments, the chamber includes a radiation aperture that is configured to allow electromagnetic waves from the heating source to pass through to the heating / cooling layer. In certain embodiments, a window is located at the radiation aperture to allow the electromagnetic waves to pass through. In certain embodiments, a filter (e.g., a bandpass filter) is positioned at the optical aperture to allow imaging of the sample in the sample holder.

[0687] In some embodiments, the chamber is used to absorb heat from the sample and / or the heating source. In some embodiments, the chamber includes a metal housing. In some embodiments, the chamber includes an outer layer. In certain embodiments, the outer layer is black. In some embodiments, the outer layer is made of black metal. In some embodiments, the chamber includes an inner layer. In some embodiments, the inner layer is made of a non-reflective material. In certain embodiments, the inner layer is black. In some embodiments, the inner layer is made of black metal.

[0688] As shown in FIGS. 13 and 14, in some embodiments, the system includes an optical sensor that is configured to capture an image of a fluid sample in the sample holder. In some embodiments, the system further includes a light source that can be integrated with the optical sensor in some cases and separated in some cases. In some embodiments, the light source is configured to provide excitation light that can reach the sample. In some embodiments, the sample can provide signal light that can be captured by the optical sensor for imaging.

[0689] As shown in FIG. 13, in some embodiments, the heating source includes an LED or a laser diode. In certain embodiments, the heating source further includes an optical fiber coupler and an optical fiber that guides light from the LED / laser diode to the sample holder.

[0690] FIG. 14 shows a cross-sectional view of an exemplary embodiment of the present invention demonstrating a system for rapidly changing the temperature of a sample. FIG. 14 shows the detailed elements of a heating source according to one embodiment. As shown in FIG. 14, in some embodiments, the heating source includes an LED or a laser diode. In certain embodiments, the heating source further includes one or more focusing lenses that focus the electromagnetic waves from the heating source onto the sample in the sample holder.

[0691] As shown in FIG. 7, the thermal control unit 200 includes a beam expander 208 that is configured to expand the electromagnetic waves from the heating source 202 from a smaller diameter to a larger diameter. In some embodiments, the electromagnetic waves projected from the heating source 202 are sufficient to cover the entire sample contact area; however, in some embodiments, it is necessary to expand the coverage area of the electromagnetic waves projected from the heating source 202 to produce an expanded electromagnetic wave 210 that provides a heat source for all sample contact areas. The beam expander 208 employs any known technique, including but not limited to the beam expanders described in U.S. Patent Nos. 4,545,677, 4,214,813, 4,127,828, and 4,016,504 and U.S. Patent Publication Nos. 2008 / 0297912 and 2010 / 0214659, which are hereby incorporated by reference in their entireties for all purposes.

[0692] Smartphone

[0693] In some embodiments, the sample card is imaged by a mobile device. In certain embodiments, the mobile device is a smartphone, which may be used as an example.

[0694] In some embodiments, the smartphone includes a camera that can be used to image the sample in the sample card. In some embodiments, an adapter is used to hold the sample card, and the adapter is configured to be attached to the smartphone such that the sample card (and the sample therein) can be placed within the field of view of the camera.

[0695] In some embodiments, the smartphone can also be used as a control unit configured to control the device. For example, the heating and / or cooling of the sample card can be controlled using the smartphone. In certain embodiments, the smartphone is connected to the heating source and controls the electromagnetic waves from the heating source. In some embodiments, the smartphone controls the presence or absence, intensity, wavelength, frequency, and / or angle of the electromagnetic waves. In certain embodiments, the smartphone receives temperature data from a thermometer that measures the sample temperature. In certain embodiments, the smartphone controls the electromagnetic waves based on the temperature data.

[0696] In some embodiments, the smartphone can also be used as a data processing and communication device. For example, after imaging a sample, the image can be saved in the smartphone. In certain embodiments, saving the image can be processed by software or an application in the smartphone. For example, the presence and / or amount of an analyte can be inferred from the image by software or an application in the smartphone. In certain embodiments, the processing result can be displayed on the screen of the smartphone. In certain embodiments, the processing result can be sent to the user, for example, using email or other messaging software. In certain embodiments, the processing result can be sent to a third party, such as a healthcare professional, who can perform further diagnosis and / or process the data in an additional step. In some embodiments, the unprocessed image can be displayed and / or sent. In certain embodiments, the image is displayed on the screen of the smartphone. In certain embodiments, the image is sent to the user, for example, by email or other messaging software. In certain embodiments, the image can be sent to a third party, such as a remote server, which can further process the image. In some embodiments, the result and / or image is compressed and / or encrypted before being sent.

[0697] Use of the RHC card

[0698] The RHC card in the specification can be used as one step in multiple steps of testing a sample, or as one step in performing the entire test.

[0699] In some embodiments, the RHC card is used for a so-called "one-step assay", in which all reagents and samples for analysis are loaded on the RHC card, a thermal cycle or temperature change is performed, and signals are observed during the thermal cycle or temperature change.

[0700] Other embodiments

[0701] Example - 1:

[0702] One embodiment includes the devices of Examples SH-1 to SH-6, wherein the first and second plates are flexible plastic films and / or thin glass films, and each plate has a substantially uniform thickness with a value selected from the range of 1 μm to 25 μm.

[0703] The area of each plate is 1 cm 2 to 16 cm 2 .

[0704] The sample sandwiched between the two plates has a thickness of 40 μm or less.

[0705] The relevant sample to whole sample ratio (RE ratio) is 12% or less.

[0706] The cooling zone is at least 9 times larger than the heating zone.

[0707] The thermal mass ratio of the sample to the non-sample is 2.2 or greater.

[0708] The RHC does not have a spacer in some embodiments, but has a spacer in other embodiments.

[0709] The STC ratio is and the cooling zone contains a material layer with a thermal conductivity of 70 W / m-K or higher and the thermal conductivity multiplied by its thickness.

[0710] Example - 2:

[0711] For the embodiments of SH-1 to SH-x, they have the following parameters arranged for rapid thermal cycling.

[0712] The first and second plates are plastic or thin glass. The thickness of the first and second plates is 100 nm, 500 nm, 1 μm, 5 μm, 10 μm, or within the range between any two values.

[0713] The thickness of the sample between the two plates is 5 μm, 10 μm, 30 μm, 50 μm, 100 μm, or within the range between any two values.

[0714] The distance from the H / C layer to the sample is 10 nm, 100 nm, 500 nm, 1 μm, 5 μm, 10 μm, or within the range between any two values.

[0715] The ratio of the cooling zone area to the related sample area is 16, 9, 4, 2, or within the range between any two values.

[0716] The ratio of the cooling zone area to the heating zone area is 16, 9, 4, 2, or within the range between any two values.

[0717] The distance between the H / C layer and the heating source (e.g., LED) is 5 mm, 10 mm, 20 mm, 30 mm, or within the range between any two values.

[0718] Example - 3:

[0719] For the embodiments of SH-1 to SH-x, they have the following parameters arranged for rapid thermal cycling.

[0720] The first and second plates are plastic or thin glass. The thickness of the first plate is 10 μm, 25 μm, 50 μm, or within the range between any two values; while the thickness of the second plate (the plate with the heating or cooling layer) is 100 nm, 500 nm, 1 μm, 5 μm, 10 μm, or within the range between any two values.

[0721] The thickness of the sample between the two plates is 5μm, 10μm, 30μm, 50μm, 100μm, or within the range between any two values.

[0722] The distance between the H / C layer and the sample is 10nm, 100nm, 500nm, 1μm, 5μm, 10μm, or within the range between any two values.

[0723] The ratio of the cooling zone area to the area of the relevant sample is 16, 9, 4, 2, or within the range between any two values.

[0724] The ratio of the cooling zone area to the heating zone area is 16, 9, 4, 2, or within the range between any two values.

[0725] The distance between the H / C layer and the heat source (e.g., LED) is 5mm, 10mm, 20mm, 30mm, or within the range between any two values.

[0726] Example - 4:

[0727] For the examples of SH - 1 to SH - x, they have the following parameters arranged for rapid thermal cycling.

[0728] The first and second plates are plastic or thin glass. The thickness of the first and second plates is 100nm, 500nm, 1μm, 5μm, 10μm, 25μm, 50μm, 100μm, 175μm, 250μm, or within the range between any two values.

[0729] The thickness of the sample between the two plates is 100nm, 500nm, 1μm, 5μm, 10μm, 25μm, 50μm, 100μm, 175μm, 250μm, or within the range between any two values.

[0730] The distance between the H / C layer and the sample is 100nm, 500nm, 1μm, 5μm, 10μm, 25μm, 50μm, 100μm, 175μm, 250μm, or within the range between any two values.

[0731] The ratio of the cooling zone area to the area of the relevant sample is 100, 64, 16, 9, 4, 2, 1, 0.5, 0.1, or within the range between any two values.

[0732] The ratio of the cooling zone area to the heating zone area is 100, 64, 16, 9, 4, 2, 1, 0.5, 0.1, or within the range between any two values.

[0733] The distance between the H / C layer and the heating source (e.g., LED) is 500 μm, 1 mm, 3 mm, 5 mm, 10 mm, 20 mm, 30 mm, or within the range between any two values.

[0734] Example - 5:

[0735] For the examples of SH - 1 to SH - 5, they have the following parameters arranged for rapid thermal cycling.

[0736] The light guide calibrates the light from the light source (e.g., LED) into the heating zone. The light stack contains a structure (e.g., a tube or a milled - hole structure) with a hollow hole having a reflective wall. The light stack has a lateral dimension of 1 mm to 8 mm and a length of 2 mm to 50 mm.

[0737] Example - 6:

[0738] For the examples of SH - 1 to SH - 5, they have the following parameters arranged for rapid thermal cycling.

[0739] The first plate and the second plate are plastic or thin glass. The thickness of the first plate and the second plate is 100 nm, 500 nm, 1 μm, 5 μm, 10 μm, or within the range between any two values.

[0740] The thickness of the sample between the two plates is 1 to 5 μm, 5 μm to 10 μm, 10 to 30 μm, or 30 μm to 50 μm.

[0741] The distance from the H / C layer to the sample is within the range of 10 nm to 100 nm, 100 nm to 500 nm, 500 nm to 1 μm, 1 μm to 5 μm, 5 μm to 10 μm, or 10 μm to 25 μm.

[0742] The ratio of the cooling - zone area to the related sample area is 16, 9, 4, 2, or within the range between any two values.

[0743] The ratio of the cooling - zone area to the heating - zone area is 16, 9, 4, 2, or within the range between any two values.

[0744] The distance between the H / C layer and the heating source (e.g., LED) is 5 mm, 10 mm, 20 mm, 30 mm, or within the range between any two values.

[0745] The KC ratio of the cooling layer is between 0.5 cm 2 / sec and 0.7 cm 2 / sec, 0.7 cm 2 / sec and 0.9 cm 2 / sec, 0.9 cm 2 / sec and 1 cm2 / sec, 1 cm 2 / sec and 1.1 cm 2 / sec, 1.1 cm 2 / sec and 1.3 cm 2 / sec, 1.3 cm 2 / sec and 1.6 cm 2 / sec, 1.6 cm 2 / sec and 2 cm 2 / sec, or 2 cm 2 / sec and 3 cm 2 within the range between / sec and 3 cm

[0746] The thermal mass ratio of the sample to the non - sample is in the range of 0.2 to 0.5, 0.5 to 0.7, 0.7 to 1, 1 to 1.5, 1.5 to 5, 5 to 10, 10 to 30, 30 to 50, or 50 to 100.

[0747] Example - 7:

[0748] For the examples of SH - 1 to SH - 5 and Examples 1 to 6, they have the following parameters arranged for rapid thermal cycling:

[0749] The lateral area of the first plate and / or the second plate is 1 mm 2 (square millimeters) to 10 mm 2 、10 mm 2 to 50 mm 2 、50 mm 2 to 100 mm 2 、1 cm 2 to 5 cm 2 、5 cm 2 to 20 cm 2 or 20 cm 2 to 50 cm 2 .

[0750] The scaled thermal conductivity (STM ratio) is in the range of 10 to 20, 30 to 50, 50 to 70, 70 to 100, 100 to 1000, 1000 to 10000, or 10000 to 1000000; and the thermal conductivity of the cooling zone (layer) multiplied by its thickness is 6×10 -5 W / K, 9×10 -5 W / K, 1.2×10 -4 W / K, 1.5×10 -4 W / K, 1.8×10 -4 W / K, 2.1×10 -4 W / K, 2.7×10 -4 W / K, 3×10 -4W / K, 1.5×10 -4 W / K, or within a range between any two values.

[0751] The sample holder (RHC card) has no significant heat conduction to the environment during thermal cycling.

[0752] Sample type

[0753] The devices, systems, and methods disclosed herein can be used for samples, such as but not limited to diagnostic samples, clinical samples, environmental samples, and food samples. The types of samples include but are not limited to the samples listed, described, and summarized in PCT Application No. PCT / US2016 / 045437 (designating the United States) filed on August 10, 2016, and PCT Application No. PCT / US0216 / 051775 (designating the United States) filed on September 14, 2016, the entire contents of which are incorporated herein by reference.

[0754] For example, in some embodiments, the devices, systems, and methods disclosed herein are used for samples including cells, tissues, body fluids, and / or mixtures thereof. In some embodiments, the sample comprises a human body fluid. In some embodiments, the sample comprises at least one of cells, tissues, body fluids, feces, amniotic fluid, aqueous humor, vitreous humor, blood, whole blood, fractionated blood, plasma, serum, breast milk, cerebrospinal fluid, earwax, chyle, chyme, endolymph, perilymph, feces, gastric acid, gastric juice, lymph, mucus, nasal drainage, sputum, pericardial fluid, peritoneal fluid, pleural fluid, pus, mucosal secretions, saliva, sebum, semen, sputum, sweat, synovial fluid, tears, vomit, urine, and exhaled breath condensate.

[0755] In some embodiments, the devices, systems, and methods disclosed herein are used for environmental samples obtained from any suitable source, such as but not limited to: rivers, lakes, ponds, oceans, glaciers, icebergs, rain, snow, sewage, reservoirs, tap water, drinking water, etc.; the solid samples are from soil, compost, sand, rock, concrete, wood, brick, sewage, etc.; and gas samples, the gas samples are from air, underwater vents, industrial exhaust gases, vehicle exhaust gases, etc. In certain embodiments, the environmental sample source is fresh; in certain embodiments, the environmental sample is processed. For example, before applying the devices, systems, and methods, the sample in non - liquid form is converted into liquid form.

[0756] In some embodiments, the devices, systems, and methods disclosed herein are for food samples that are suitable or may become suitable for animal consumption, such as human consumption. In some embodiments, food samples include raw materials, cooked or processed foods, foods of plant and animal origin, pre-processed foods, and partially or fully processed foods, among others. In certain embodiments, the sample in non-liquid form is converted into a liquid form prior to applying the devices, systems, and methods.

[0757] The devices, systems, and methods can be used to analyze samples of any volume. Examples of volumes include, but are not limited to, about 10 mL or less, 5 mL or less, 3 mL or less, 1 microliter (μL, also referred to herein as "μL") or less, 500 μL or less, 300 μL or less, 250 μL or less, 200 μL or less, 170 μL or less, 150 μL or less, 125 μL or less, 100 μL or less, 75 μL or less, 50 μL or less, 25 μL or less, 20 μL or less, 15 μL or less, 10 μL or less, 5 μL or less, 3 μL or less, 1 μL or less, 0.5 μL or less, 0.1 μL or less, 0.05 μL or less, 0.001 μL or less, 0.0005 μL or less, 0.0001 μL or less, 10 pL or less, 1 pL or less, or a range between any two of these values.

[0758] In some embodiments, the volume of the sample includes, but is not limited to, about 100 μL or less, 75 μL or less, 50 μL or less, 25 μL or less, 20 μL or less, 15 μL or less, 10 μL or less, 5 μL or less, 3 μL or less, 1 μL or less, 0.5 μL or less, 0.1 μL or less, 0.05 μL or less, 0.001 μL or less, 0.0005 μL or less, 0.0001 μL or less, 10 pL or less, 1 pL or less, or a range between any two of these values. In some embodiments, the volume of the sample includes, but is not limited to, about 10 μL or less, 5 μL or less, 3 μL or less, 1 μL or less, 0.5 μL or less, 0.1 μL or less, 0.05 μL or less, 0.001 μL or less, 0.0005 μL or less, 0.0001 μL or less, 10 pL or less, 1 pL or less, or a range between any two of these values.

[0759] In some embodiments, the amount of the sample is about one drop of liquid. In certain embodiments, the amount of the sample refers to the amount collected from a pricked finger or finger stick. In certain embodiments, the amount of the sample refers to the amount collected from a microneedle, micropipette, or venipuncture.

[0760] In some embodiments, the sample holder is configured to hold a fluid sample. In some embodiments, the sample holder is configured to squeeze at least a portion of the fluid sample into a thin layer. In some embodiments, the sample holder includes structures configured to heat and / or cool the sample. In some embodiments, the heat source provides electromagnetic waves that can be absorbed by certain structures in the sample holder to change the temperature of the sample. In some embodiments, the signal sensor is configured to detect and / or measure a signal from the sample. In some embodiments, the signal sensor is configured to detect and / or measure an analyte in the sample. In some embodiments, the heat sink is configured to absorb heat from the sample holder and / or the heat source. In some embodiments, the heat sink includes a chamber that at least partially surrounds the sample holder.

[0761] Applications

[0762] The devices, systems, and methods disclosed herein can be used for various types of biological / chemical sampling, sensing, assays, and applications, including those listed, described, and summarized in PCT Application No. PCT / US2016 / 045437, filed Aug. 10, 2016 (designating the United States), which is hereby incorporated by reference in its entirety.

[0763] In some embodiments, the devices, systems, and methods disclosed herein are used in a variety of different applications in various fields where the presence, quantification, and / or amplification of one or more analytes in a sample needs to be detected. For example, in certain embodiments, the devices, systems, and methods are used to detect proteins, peptides, nucleic acids, synthetic compounds, inorganic compounds, organic compounds, bacteria, viruses, cells, tissues, nanoparticles, and other molecules, compounds, mixtures, and substances. Various fields in which the devices, systems, and methods can be used include, but are not limited to: diagnosis, management, and / or prevention of human diseases and conditions, diagnosis, management, and / or prevention of animal diseases and conditions, diagnosis, management, and / or prevention of plant diseases and conditions, agricultural uses, veterinary uses, food testing, environmental testing and remediation, drug testing and prevention, etc.

[0764] The applications of the present invention include, but are not limited to: (a) detecting, purifying, quantifying, and / or amplifying compounds or biomolecules associated with certain diseases (such as infectious and parasitic diseases, injuries, cardiovascular diseases, cancers, mental disorders, neuropsychiatric disorders, and organic diseases (such as lung diseases, kidney diseases)) or certain stages of these diseases, (b) detecting, purifying, quantifying, and / or amplifying cells and / or microorganisms (such as viruses, fungi, and bacteria) from the environment (such as water, soil, or biological samples (such as tissues, body fluids)), (c) detecting and quantifying compounds or biological samples (such as toxic waste, anthrax) that pose a hazard to food safety, human health, or national security, (d) detecting and quantifying vital parameters (such as glucose, blood oxygen level, total blood cell count) in medical or physiological monitors, (e) detecting and quantifying specific DNA or RNA from biological samples (such as cells, viruses, body fluids), (f) performing sequencing and comparison of genetic sequences of DNA in chromosomes and mitochondria for genomic analysis, or (g) detecting and quantifying reaction products, for example, during the synthesis or purification of drugs.

[0765] In some embodiments, the devices, systems, and methods are used to detect nucleic acids, proteins, or other molecules or compounds in a sample. In certain embodiments, the devices, systems, and methods are used for rapid clinical detection and / or quantification of one or more, two or more, or three or more disease biomarkers in a biological sample, for example, for diagnosing, preventing, and / or managing a disease condition in a subject. In certain embodiments, the devices, systems, and methods are used to detect and / or quantify one or more, two or more, or three or more environmental markers in an environmental sample, which is a sample obtained, for example, from a river, ocean, lake, rain, snow, sewage, sewage treatment runoff, agricultural runoff, industrial runoff, tap water, or drinking water. In certain embodiments, the devices, systems, and methods are used to detect and / or quantify one or more, two or more, or three or more food markers from a food sample, which is obtained from tap water, drinking water, prepared food, processed food, or unprocessed food.

[0766] In some embodiments, the devices, systems, and methods of the present invention can be used to detect analytes. In some embodiments, the analyte is a pathogen. Example pathogens that can be detected include, but are not limited to: varicella zoster; Staphylococcus epidermidis, Escherichia coli, methicillin-resistant Staphylococcus aureus (MSRA), Staphylococcus aureus, Staphylococcus hominis, Enterococcus faecalis, Pseudomonas aeruginosa, Staphylococcus capitis, Staphylococcus warneri, Klebsiella pneumoniae, Haemophilus influenzae, Staphylococcus simulans, Streptococcus pneumoniae, and Candida albicans; gonorrhea (Neisseria gonorrhoeae), syphilis (Treponema pallidum), chlamydia (Chlamydia trachomatis), non-gonococcal urethritis (Ureaplasm urealyticum), chancroid (Haemophilus ducreyi), trichomonas (trichomonas vaginalis); Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus (MSRA), Klebsiella pneumoniae, Haemophilus influenzae, Staphylococcus aureus, Stenotrophomonas maltophilia, Haemophilus parainfluenzae, Escherichia coli, Enterococcus faecalis, Serratia, Haemophilus parahaemolyticus, Enterococcus cloacae, Candida albicans, Moraxella catarrhalis, Streptococcus pneumoniae, Citrobacter freundii, Enterococcus faecalis, Klebsiella oxytoca, Pseudomonas fluorescens, Neisseria meningitidis, Streptococcus pyogenes, Pneumocystis carinii, Klebsiella pneumoniae, Legionella pneumophila, Mycoplasma pneumoniae, and Mycobacterium tuberculosis, etc.

[0767] In some embodiments, the devices, systems, and methods of the present invention can be used to detect analytes that are diagnostic markers. In some embodiments, the diagnostic markers are selected from any of the following tables.

[0768] Table 4.1: Diagnostic Markers

[0769]

[0770]

[0771] Table 4.2: Diagnostic Markers

[0772]

[0773]

[0774]

[0775]

[0776]

[0777]

[0778]

[0779]

[0780]

[0781]

[0782]

[0783]

[0784]

[0785]

[0786]

[0787]

[0788]

[0789] In some embodiments, the devices, systems, and methods of the present invention can be used to inform a subject from whom a sample is obtained of their health status. Health conditions that can be diagnosed or measured by the methods, devices, and systems include, but are not limited to: chemical balance; nutraceuticals; exercise; fatigue; sleep; stress; prediabetes; allergies; aging; exposure to environmental toxins, pesticides, herbicides, synthetic hormone analogs; pregnancy; menopause; and male menopause. Table 4.3 provides exemplary diagnostic markers that can be detected using the present invention and their associated health conditions.

[0790] Table 4.3: Diagnostic Markers

[0791]

[0792]

[0793]

[0794]

[0795]

[0796]

[0797]

[0798]

[0799] In other embodiments, diagnostic markers detectable by the present method are antibodies in a sample (e.g., a diagnostic sample) that can be used to diagnose a disease or health condition of a subject from whom the sample was obtained. Table 4.4 provides a list of 5 autoantibody targets that can be used in whole or as epitope fragments as capture agents in the present method to measure the amount of epitope-binding antibody analyte in a sample, thereby diagnosing related diseases or health conditions, such as autoimmune diseases. In some cases, the disease or health condition is related to an immune response to an allergen. Table 4.5 provides a list of allergens that can be used in whole or as epitope fragments as capture agents in the present method to measure the amount of epitope-binding antibody analyte in a sample, thereby diagnosing related diseases or health conditions, such as allergies. In certain cases, the disease or health condition is related to an infectious disease, where the infectious agent can be diagnosed based on information including the measured amount of antibodies against one or more epitopes from the infectious agent (e.g., lipopolysaccharides, toxins, proteins, etc.). Table 4.6 provides a list of infectious agent-derived epitopes that can be used in whole or as epitope fragments as capture agents in the method of the present invention to measure the amount of epitope-binding antibody analyte in a sample and thereby diagnose related diseases or health conditions (e.g., infections). Other epitopes or antigens suitable for the present diagnostic method are described, for example, in PCT application publication No. WO2013164476, which is incorporated herein by reference.

[0800] Table 4.4: Diagnostic Autoantibody Epitopes

[0801]

[0802]

[0803]

[0804]

[0805]

[0806]

[0807] Table 4.5: Allergen Epitopes

[0808]

[0809]

[0810]

[0811]

[0812] Table 4.6: Infectious Agent-Derived Epitopes

[0813]

[0814]

[0815] In some embodiments, the devices, systems, and methods of the present invention can be used to detect diagnostic markers, which are microRNA (miRNA) biomarkers associated with diseases or health conditions. Table 4.7 provides an exemplary list of miRNA biomarkers that can be used and their associated diseases / health conditions.

[0816] Table 4.7: Diagnostic miRNA Markers

[0817]

[0818]

[0819]

[0820]

[0821]

[0822]

[0823] *miRNA markers in parentheses are downregulated

[0824] In some embodiments, the devices, systems, and methods of the present invention can be used to detect or analyze environmental samples. Environmental samples can be obtained from any suitable source, such as rivers, oceans, lakes, rain, snow, sewage, sewage treatment runoff, agricultural runoff, industrial runoff, water, tap water, or drinking water, etc.).

[0825] In some embodiments, the analyte that can be detected or analyzed using the devices, systems, and methods of the present invention is an environmental marker. An environmental marker can be any suitable marker that can be captured by a capture agent, which specifically binds the environmental marker in a device configured with the capture agent. In some embodiments, the devices, systems, and methods of the present invention detect the concentration of lead or toxins in water. In some embodiments, the presence, absence, or quantitative level of an environmental marker in a sample can indicate the state of the environment from which the sample was obtained. In some embodiments, an environmental marker can be a substance that is toxic or harmful to organisms exposed to the environment, such as humans, companion animals, plants, etc. In some embodiments, an environmental marker can be an allergen, which can cause an allergic reaction in some individuals exposed to the environment. In some embodiments, the presence, absence, or quantitative level of an environmental marker in a sample can be related to the general health of the environment. In this case, the overall health of the environment can be measured over a period of time (e.g., one week, several months, several years, or several decades).

[0826] In some embodiments, the devices, systems, and methods of the present invention further include receiving or providing a report that indicates the safety or harmfulness of a subject to be exposed to the environment from which a sample was obtained, based on information including the amount of an environmental marker that was measured. Information for assessing the safety risk or health of an environment can include data in addition to the type and measured amount of the environmental marker. Such other data can include, for example, location, altitude, temperature, time of day / month / year, pressure, humidity, wind direction and speed, weather, and the like. The data can represent, for example, an average value or trend over a particular time period (minutes, hours, days, weeks, months, years, etc.) or an instantaneous value over a shorter time period (milliseconds, seconds, minutes, etc.).

[0827] In some embodiments, the report can be generated by a device configured to read the device, or can be generated at a remote location when data including the amount of the measured environmental marker is sent. In some embodiments, an expert can be at a remote location or can obtain the data sent to a remote location and can analyze or review the data to generate a report. In some embodiments, the expert can be a scientist or administrator from a government agency (such as the U.S. Centers for Disease Control and Prevention (CDC) or the U.S. Environmental Protection Agency (EPA)), a research institution (such as a university), or a private company. In some embodiments, the expert can send instructions or recommendations to a user based on the data sent by the device and / or analyzed at a remote location.

[0828] A list of exemplary environmental markers is set forth in Table 8 of U.S. Provisional Application Serial No. 62 / 234,538, filed on September 29, 2015, which is incorporated herein by reference.

[0829] Other exemplary environmental markers are listed in Table 4.8.

[0830] Table 4.8: Environmental Markers

[0831]

[0832]

[0833]

[0834]

[0835]

[0836] In some embodiments, the devices, systems, and methods of the present invention can be used to detect or analyze food samples. The food samples can be obtained from any suitable source, such as raw food, processed food, cooked food, drinking water, etc. In some embodiments, the analyte that can be detected or analyzed using the devices, systems, and methods of the present invention is a food marker. The food marker can be, for example, any suitable marker shown in Table 4.9 that can be captured by a capture agent that specifically binds the food marker in a device configured with the capture agent. In some embodiments, the presence, absence, or quantitative level of the food marker in the sample can indicate the safety or hazard of the subject if the food is consumed. In some embodiments, the food marker is a substance derived from a pathogen or microorganism that indicates the presence or absence of the organism in the food from which the sample was obtained. In some embodiments, the food marker is a toxic or harmful substance if consumed by the subject. In some embodiments, the food marker is a bioactive compound that may inadvertently or accidentally alter physiology if consumed by the subject.

[0837] In some embodiments, the food marker is an indication of the manner in which the food was obtained (e.g., grown, acquired, caught, harvested, processed, cooked, etc.). In some embodiments, the food marker indicates the nutritional content of the food. In some embodiments, the food marker is an allergen that can induce an allergic reaction if the food from which the sample was obtained is consumed by the subject.

[0838] In some embodiments, the devices, systems, and methods of the present invention further include receiving or providing a report that indicates the safety or hazard of the subject consuming the food from which the sample was obtained based on information including the measured level of the food marker. Information for evaluating the safety of food for consumption can include data of types and measured amounts different from the food marker. These other data can include any health conditions related to the consumer (allergies, pregnancy, chronic or acute diseases, current prescription medications, etc.).

[0839] The report can be generated by a device configured to read the device or can be generated at a remote location when data including the measured amount of the food marker is sent. In some cases, a food safety expert can be at a remote location or can access the data sent to a remote location and can analyze or view the data to generate a report. In some embodiments, the food safety expert can be a scientist or administrator of a government agency (e.g., the U.S. Food and Drug Administration (FDA) or the CDC), a research institution (e.g., a university), or a private company. In certain embodiments, the food safety expert can send instructions or recommendations to a user based on the data sent by the device and / or analyzed at a remote location.

[0840] Exemplary food markers are listed in Table 9 of U.S. Provisional Application Serial No. 62 / 234,538, filed September 29, 2015, which is incorporated herein by reference.

[0841] Other exemplary food markers are listed in Table 4.9.

[0842] Table 4.9: Food Markers

[0843]

[0844]

[0845]

[0846] In some embodiments, the present invention relates to a kit comprising the device of the present invention. In some embodiments, the kit includes a device configured to specifically bind an analyte as described herein. In some embodiments, the kit includes instructions for practicing the methods of the present invention using a handheld device such as a mobile phone. In some embodiments, the instructions can be present in the kit in a variety of forms, one or more of which can be present in the kit. One form in which these instructions can be present is printed information on a suitable medium or substrate (such as one or more sheets of paper on which information is printed, in the packaging of the kit, in a package insert, etc.). Another device can be a computer-readable medium on which information is recorded or stored, such as a disk, CD, DVD, Blu-ray, computer-readable memory, etc. Another device that can be present is a website address, which can be used via the Internet to access information at a removed site. The kit can further include software for practicing a method for measuring an analyte on a device as provided on a computer-readable medium. Any convenient device can be present in the kit.

[0847] In some embodiments, the kit includes a detection reagent containing a detectable label (such as a fluorescently labeled antibody or oligonucleotide that specifically binds a relevant analyte) for labeling the relevant analyte. The detection agent can be provided as a device in a separate container, or can be provided in the device.

[0848] In some embodiments, the kit includes a control sample that includes a known detectable amount of the analyte to be detected in a sample. The control sample can be provided in a container, and can be provided in a solution of known concentration, or can be provided in a dry form (such as lyophilized or freeze-dried). If provided in a dry form, the kit can further include a buffer for dissolving the control sample.

[0849] In some embodiments, the devices, systems, and methods of the present invention can be used for simple and rapid blood cell counting using a smartphone. In some embodiments, the first and second plates are selected from thin glass slides (e.g., 0.2 mm thick) or thin plastic films (e.g., 15 mm thick) with relatively flat surfaces, and each has an area with a length and width of about 0.5 cm to 10 cm. In some embodiments, the spacer is made of glass, plastic, or other materials that do not deform significantly under pressure. In some embodiments, the spacer is placed on the first plate, the second plate, or both before sample deposition; and the first plate, the second plate, or both are optionally coated with reagents (staining dyes and / or anticoagulants) that facilitate blood counting. In some embodiments, the first and second plates can be sealed in a bag for ease of transportation and to achieve a longer shelf life.

[0850] In some embodiments of the blood cell counting test, the sample only requires about 1 μL (microliter) (or about 0.1 μL to 3 μL) of blood that can be taken from, for example, a finger or other human body part. In some embodiments, the blood sample can be directly deposited onto the first and second plates from the human body (e.g., a finger) without any dilution. In such embodiments, the first and second plates can face each other such that the blood sample is located between the inner surfaces of the first and second plates. In some embodiments, the reagents are pre-deposited (staining dyes or anticoagulants), and they are deposited on the inner surfaces to mix with the sample. Then, the first and second plates can be pressed by a finger or a simple mechanical device (e.g., a clip pressed using a spring). Under pressure, the internal spacing is reduced and finally terminates at a value set by the height of the spacer, and the final sample thickness is reached, which is typically equal to the final internal spacing. Since the final internal spacing is known, the final sample thickness becomes known, i.e., it is quantified (measured) by this method.

[0851] In some embodiments, if the blood sample is not diluted, after pressing the spacer (sample deformation), and thus the final sample thickness can be thin, such as less than 1 μm, less than 2 μm, less than 3 μm, less than 4 μm, less than 5 μm, less than 7 μm, less than 10 μm, less than 15 μm, less than 20 μm, less than 30 μm, less than 40 μm, less than 50 μm, less than 60 μm, less than 80 μm, less than 100 μm, less than 150 μm, or within any range between any two values. A thin final sample may be useful because if the final sample is thick, then many red blood cells may overlap during imaging, which may make cell counting inaccurate. For example, undiluted whole blood with a thickness of about 4 μm will produce approximately one layer of red blood cells.

[0852] After pressing, the sample can be imaged directly through a smartphone or through additional optical elements (such as lenses, filters, or light sources as needed). The image of the sample can be processed to identify the type of cells and the number of cells. The image processing can be done locally on the same smartphone where the image is acquired or remotely, but the final result is transmitted back to the smartphone (where the image is transmitted to a remote location and processed there). The smartphone will display the number of cells of a particular type. In some cases, certain suggestions will be displayed. The suggestions can be stored on the smartphone prior to the test, or the suggestions can come from a remote machine or a professional.

[0853] In some embodiments, a reagent is placed on the inner surface of the first plate and / or the second plate using the methods and devices described herein.

[0854] In some embodiments, a device or method for a blood test includes (a) the device or method described herein and (b) a plate spacing in a closed configuration (i.e., the distance between the inner surfaces of two plates) or using such a spacing, wherein the lateral average intercellular distance of red blood cells (RBCs) in undiluted whole blood in the plate spacing is greater than the average diameter of the disc shape of the RBCs.

[0855] In some embodiments, a device or method for arranging the orientation of non-spherical cells includes (a) the device or method described herein, and (b) a plate spacing in a closed configuration or using such a spacing (i.e., the distance between the inner surfaces of the two plates), wherein the spacing is less than the average size of the unit in its long direction (the long direction is the direction of the maximum dimension of the unit). Such an arrangement can improve the measurement of the sample volume (such as the red blood cell volume).

[0856] In some embodiments, the analyte in a blood test includes protein markers whose list can be found on the website of the American Association for Clinical Chemistry.

[0857] Table 4.10 provides other exemplary analytes that can be detected using the present invention at the point of care (POC) setting and / or when used by a non-professional user / subject.

[0858] Table 4.10: POC Analytes

[0859]

[0860]

[0861]

[0862]

[0863]

[0864]

[0865] In some embodiments, the devices, systems, and methods of the present invention can be used to detect or diagnose a health condition. In some embodiments, the health condition includes, but is not limited to: chemical balance; nutraceuticals; exercise; fatigue; sleep; stress; prediabetes; allergies; aging; exposure to environmental toxins, pesticides, herbicides, synthetic hormone analogs; pregnancy; menopause; and male menopause.

[0866] In some embodiments, these methods can be used to obtain and compare the relative levels of nucleic acids in two or more different nucleic acid samples. In these embodiments, the results obtained from the methods herein are typically normalized to the total amount of nucleic acid in the sample (e.g., constitutive RNA) and compared. This can be done by comparing ratios or by any other means. In certain embodiments, the nucleic acid profiles of two or more different samples can be compared to identify nucleic acids associated with a particular disease or disorder.

[0867] In some embodiments, the devices, systems, and methods of the present invention can include a) obtaining a sample, b) applying the sample to a device containing a capture agent that binds to a relevant analyte under conditions suitable for binding of the analyte in the sample to the capture agent, c) washing the device, and d) reading the device, thereby obtaining a measurement of the amount of analyte in the sample. In some embodiments, the analyte can be a biomarker, an environmental marker, or a food marker. In some cases, the sample is a liquid sample and can be a diagnostic sample (e.g., saliva, serum, blood, sputum, urine, sweat, tears, semen, or mucus); an environmental sample, taken from a river, ocean, lake, rainwater, snow, sewage, sewage treatment runoff, agricultural runoff, industrial runoff, tap water, or drinking water; or a food sample obtained from tap water, drinking water, prepared food, processed food, or raw materials. In some embodiments, the device can be disposed in a microfluidic device, and the applying step b) can include applying the sample to the microfluidic device containing the device. In some embodiments, the reading step d) can include detecting a fluorescence or luminescence signal from the device. In some embodiments, the reading step d) can include using a handheld device configured to read the device to read the device. The handheld device can be a mobile phone, such as a smartphone. In some embodiments, the device can include a labeling reagent that can bind to the analyte-capture agent complex on the device. In some embodiments, between steps c) and d), the devices, systems, and methods of the present invention can further include a step of applying a labeling reagent that binds to the analyte-capture agent complex on the device and washing the device. In any embodiment, the reading step d) can include reading an identifier of the device. The identifier can be an optical barcode, a radio frequency ID tag, or a combination thereof. In some embodiments, the devices, systems, and methods of the present invention can further include applying a control sample to a control device containing a capture agent that binds to the analyte, where the control sample includes a known detectable amount of the analyte, and reading the control device, thereby obtaining a control measurement of the known detectable amount of the analyte in the sample. In some embodiments, the sample can be a diagnostic sample obtained from a subject, the analyte can be a biomarker, and the measured amount of the analyte in the sample can be a diagnosis of a disease or disorder.

[0868] In some embodiments, the devices, systems, and methods of the present invention may further include receiving or providing a report to a subject that indicates the measured amount of a biomarker and the range of measured values of the biomarker in an individual who does not have or has a low risk of having a disease or disorder, wherein the measured amount of the biomarker relative to the range of measured values is used to diagnose the disease or disorder. In some embodiments, the devices, systems, and methods of the present invention may further include diagnosing a subject based on information including the measured amount of the biomarker in a sample. In some embodiments, the diagnosing step includes sending data containing the measured amount of the biomarker to a remote location and receiving a diagnosis based on information including the measurements from the remote location. In some embodiments, the biomarker may be selected from those listed in a table. In some embodiments, the device may comprise a plurality of capture agents each binding to a biomarker described herein, wherein the reading step d) includes obtaining a measurement of the amount of the plurality of biomarkers in the sample, and wherein the amount of the plurality of biomarkers in the sample can be used to diagnose a disease or disorder. In some embodiments, the capture agent may be an antibody epitope and the biomarker may be an antibody that binds to the antibody epitope. In some embodiments, the antibody epitope includes an allergen or a fragment thereof selected from a table. In some embodiments, the antibody epitope includes an allergen or a fragment thereof selected from a table. In some embodiments, the antibody epitope includes a biomolecule derived from an infectious agent or a fragment thereof selected from a table. In some embodiments, the device may comprise a plurality of antibody epitopes selected from a table, wherein the reading step d) includes obtaining a measurement of the amount of the plurality of epitope-binding antibodies in the sample, and wherein the amount of the plurality of epitope-binding antibodies in the sample can be used to diagnose a disease or disorder.

[0869] In some embodiments, the sample may be an environmental sample and wherein the analyte may be an environmental marker. In some embodiments, the environmental markers described herein. In some embodiments, the method may include receiving or providing a report indicating the safety or hazard to a subject exposed to the environment from which the sample was obtained. In some embodiments, the method may include sending data containing the measured amount of the environmental marker to a remote location and receiving a report indicating the safety or hazard to a subject exposed to the environment from which the sample was obtained. In any embodiment, the device may include a plurality of capture agents each binding to an environmental marker described herein, and wherein the reading step d) may include obtaining a measurement of the amount of the plurality of environmental markers in the sample.

[0870] In some embodiments, the sample can be a food sample, wherein the analyte can be a food marker, and wherein the amount of the food marker in the sample can be related to the safety of consuming the food. In some embodiments, the food marker is an example as described herein. In any embodiment, the method can include receiving or providing a report indicating the safety or hazard of the food from which the sample was obtained for consumption by the subject. In any embodiment, the method can include sending data comprising the measured amount of the food marker to a remote location and receiving a report indicating the safety or hazard of the food from which the sample was obtained for consumption by the subject. In any embodiment, the device array can include a plurality of capture agents, each of which binds to a food marker described herein, wherein the obtaining can include obtaining a measurement of the amount of a plurality of food markers in the sample, and wherein the amount of the plurality of food markers in the sample can be associated with the safety of consuming the food.

[0871] In some embodiments, the device is part of a microfluidic device. In some embodiments, the device, apparatus, system, and method are for detecting fluorescence or luminescence signals. In some embodiments, the device, system, and method include a communication device or are used in conjunction with a communication device, such as but not limited to: a mobile phone, a tablet computer, and a portable computer. In some embodiments, the device, system, and method include an identifier or are used in conjunction with an identifier (such as but not limited to an optical barcode, a radio frequency ID tag, or a combination thereof).

[0872] In some embodiments, the sample is a diagnostic sample obtained from a subject, the analyte is a biomarker, and the measured amount of the analyte in the sample is a diagnostic conclusion of a disease or condition. In some embodiments, the device, system, and method further include receiving or providing to the subject a report indicating the measured amount of the biomarker and the range of measured values of the biomarker in an individual not suffering from or at low risk of suffering from the disease or condition, wherein the measured amount of the biomarker relative to the range of measured values is a diagnostic conclusion of the disease or condition.

[0873] In some embodiments, the sample is an environmental sample, and wherein the analyte is an environmental marker. In some embodiments, the device, system, and method include receiving or providing a report indicating the safety or hazard of the environment from which the sample was obtained to which the subject was exposed. In some embodiments, the device, system, and method include sending data comprising the measured amount of the environmental marker to a remote location and receiving a report indicating the safety or hazard of the environment from which the sample was obtained to which the subject was exposed.

[0874] In some embodiments, the sample is a food sample, wherein the analyte is a food marker, and wherein the amount of the food marker in the sample is related to the safety of the consumed food. In some embodiments, the device, system, and method include receiving or providing a report that indicates the safety or hazard of the food from which the sample was obtained by the subject. In some embodiments, the device, system, and method include sending data on the measured amount of the food marker to a remote location and receiving a report that indicates the safety or hazard of the food from which the sample was obtained by the subject.

[0875] A variety of samples can be used in the assays performed using the devices, apparatuses, and systems described herein. In some embodiments, the sample contains nucleic acids. In some embodiments, the sample contains proteins. In some embodiments, the sample contains carbohydrates. The current devices, apparatuses, and systems can be used to rapidly change the temperature of the sample and stably maintain the temperature of the sample, thus providing a rapid and cost-effective method for processing the sample. In addition, a variety of applications (such as assays) can be performed using the devices, apparatuses, and systems described herein. These applications include, but are not limited to, diagnostic tests, health monitoring, environmental testing, and / or forensic identification. Such applications also include, but are not limited to, a variety of biological, chemical, and biochemical assays (such as: DNA amplification, DNA quantification, selective DNA isolation, genetic analysis, tissue typing, oncogene identification, infectious disease testing, genetic fingerprinting, and / or paternity testing).

[0876] In some embodiments, a "sample" can be any nucleic acid with or without a sample, including but not limited to human body fluids such as whole blood, plasma, serum, urine, saliva, and sweat, as well as cell cultures (mammalian, plant, bacterial, fungal). The sample can be freshly obtained or stored or processed in any desired or convenient manner, such as by dilution or addition of a buffer or other solution or solvent. Cellular structures such as human cells, animal cells, plant cells, bacterial cells, fungal cells, and virus particles can be present in the sample.

[0877] As used herein, the term "nucleic acid" refers to any DNA or RNA molecule, or DNA / RNA hybrid, or a mixture of DNA and / or RNA. Thus, the term "nucleic acid" is intended to include, but not limited to, genomic or chromosomal DNA, plasmid DNA, amplified DNA, cDNA, total RNA, mRNA, and small RNAs. The term "nucleic acid" is also intended to include natural DNA and / or RNA molecules, or synthetic DNA and / or RNA molecules. In some embodiments, cell-free nucleic acids are present in the sample. As used herein, "cell-free" means that the nucleic acid is not contained within any cellular structure. In some other embodiments, the nucleic acid is contained within a cellular structure, and the cellular structure includes, but is not limited to, the methods of the present invention are applicable to samples within a certain volume range. Samples of different volumes can be introduced onto plates of different sizes.

[0878] As used herein, "nucleic acid amplification" includes any technique for detecting nucleic acids by amplifying (producing a large number of copies of) a target molecule in a sample, where "target" herein refers to a sequence or a partial sequence of the relevant nucleic acid. Suitable nucleic acid amplification techniques include, but are not limited to, different polymerase chain reaction (PCR) methods, such as, hot start PCR, nested PCR, touchdown PCR, reverse transcription PCR, RACE PCR, digital PCR, etc., and isothermal amplification methods, such as, loop-mediated isothermal amplification (LAMP), strand displacement amplification, helicase-dependent amplification, nicking enzyme amplification, rolling circle amplification, recombinase polymerase amplification, etc.

[0879] As used herein, "essential reagents" or "reagents" include, but are not limited to, primers, deoxynucleotides (dNTPs), divalent cations (e.g., Mg 2+ ), monovalent cations (e.g., K + ), buffers, enzymes, additives, and reporters. "Essential reagents for nucleic acid amplification" are in dry form on the inner surface of the first plate or the second plate or both, or in liquid form encapsulated, embedded, or surrounded by a material (e.g., paraffin) that melts with increasing temperature.

[0880] As used herein, "primer", in some embodiments, can refer to a pair of forward and reverse primers. In some embodiments, the primer can refer to multiple primers or primer sets. As used herein, enzymes suitable for nucleic acid amplification include, but are not limited to, DNA-dependent polymerases, or RNA-dependent DNA polymerases, or DNA-dependent RNA polymerases. Examples of suitable DNA-dependent polymerases include, but are not limited to, AptaTaq polymerase, Kapa2G Fast polymerase, Kapa2G Robust, Z-Taq polymerase, Terra PCR direct polymerase, SpeedStar HS DNA polymerase, Phusion DNA polymerase, and high-fidelity DNA polymerase.

[0881] As used herein, "additive" in some embodiments includes, but is not limited to, 7-deaza-2'-deoxyguanosine 7-deaza dGTP, BSA, gelatin, betaine, DMSO, formamide, Tween 20, NP-40, Triton X-100, and tetramethylammonium chloride.

[0882] As used herein, the term "reporter" refers to any label, marker, or dye that can bind to or intercalate into a nucleic acid molecule or be activated by a byproduct of an amplification process to visualize the nucleic acid molecule or the amplification process. Suitable reporters include, but are not limited to, fluorescent labels or tags or dyes, intercalating agents, molecular beacon labels, or bioluminescent molecules, or combinations thereof.

[0883] In some other embodiments, as used herein, "essential reagent" or "reagent" (e.g., for nucleic acid amplification reactions) may also include cell lysis reagents that help break down cell structures. Cell lysis reagents include, but are not limited to, salts, detergents, enzymes, and other additives. The term "salt" as used herein includes, but is not limited to, lithium salts (e.g., lithium chloride), sodium salts (e.g., sodium chloride), and potassium salts (e.g., potassium chloride). The term "detergent" as used herein can be ionic, including anionic and cationic, non-ionic, or zwitterionic. The term "ionic detergent" as used herein includes any detergent that is partially or fully in ionic form when dissolved in water. Suitable anionic detergents include, but are not limited to, sodium dodecyl sulfate (SDS) or other alkali metal alkyl sulfates or similar detergents, Sarkosyl, or combinations thereof. The term "enzyme" as used herein includes, but is not limited to, lysozyme, cellulase, and protease. In addition, chelating agents can be included in the cell lysis reagents, including, but not limited to, EDTA, EGTA, and other polyaminocarboxylic acids, and some reducing agents, such as dithiothreitol (dTT). The composition of the essential reagents herein varies according to the rational design of different amplification reactions. In some embodiments, for example, when performing isothermal amplification by LAMP, the sample is heated to 60 - 65 °C for about 1 - 70 minutes.

[0884] As used herein, "nucleic acid amplification product" refers to various nucleic acids produced by nucleic acid amplification techniques. The types of nucleic acid amplification products herein include, but are not limited to, single-stranded DNA, single-stranded RNA, double-stranded DNA, linear DNA, or circular DNA, etc. In some embodiments, the nucleic acid amplification products can be the same nucleic acids with the same length and structure. In some other embodiments, the nucleic acid amplification products can be multiple nucleic acids with different lengths and structures.

[0885] In some embodiments, a reporter is used to quantify the nucleic acids accumulated after nucleic acid amplification. As defined and used above, the reporter has a quantifiable characteristic that is related to the presence or absence, or quantity, of nucleic acid amplicons accumulated in a closed chamber.

[0886] As used herein, "cell lysis reagent" includes, but is not limited to, salts, detergents, enzymes, and other additives that facilitate the disruption of cell structure. The term "salt" as used herein includes, but is not limited to, lithium salts (such as lithium chloride), sodium salts (such as sodium chloride), and potassium salts (such as potassium chloride). The term "detergent" as used herein can be ionic, including anionic and cationic, non-ionic, or zwitterionic. The term "ionic detergent" as used herein includes any detergent that is partially or fully in ionic form when dissolved in water. Suitable anionic detergents include, but are not limited to, sodium dodecyl sulfate (SDS) or other alkali metal alkyl sulfates or similar detergents, Sarkosyl, or combinations thereof. The term "enzyme" as used herein includes, but is not limited to, lysozyme, cellulase, and protease. In addition, chelating agents can be included in the cell lysis reagent, including, but not limited to, EDTA, EGTA, and other polyaminocarboxylic acids, and some reducing agents, such as dithiothreitol (dTT). The composition of the essential reagents herein varies according to the rational design of different amplification reactions.

[0887] As used herein, "essential reagent 2" includes, but is not limited to, primers, deoxynucleotides (dNTPs), divalent cations (such as Mg 2+ 2+ + +

[0888] ), monovalent cations (such as K

[0889] ), buffers, enzymes, and reporters. The essential reagent 2 for nucleic acid amplification can be in dry form on the inner surface of the first plate or the second plate or both, or in liquid form encapsulated, embedded, or surrounded by a material that melts with increasing temperature (such as paraffin).

[0890] The terms "CROF card (or card)", "COF card", "QMAX card", "Q card", "CROF device", "COF device", "QMAX device", "CROF plate", "COF plate", and "QMAX plate" are interchangeable and can be used to identify embodiments of the devices described herein.

[0891] The term "X plate" refers to one of two plates in a CROF card to which a spacer is fixed. The COF card, CROF card, and X plate are described in more detail in provisional application serial number 62 / 456065 filed on February 7, 2017, the entire content of which is incorporated herein by reference for all purposes.

[0892] The RHC card is a QMAX card that has or does not have a spacer plus a heating / cooling layer above or within one of its plates.

[0893] Figure 5 shows a device card 100 that includes a first plate 10 and a second plate 20. In some embodiments, the first plate 10 and the second plate 20 can be moved relative to each other into different configurations, including an open configuration and a closed configuration. In certain embodiments, in the open configuration, the two plates are partially or fully separated, and the average spacing between the plates is at least 300 μm. In certain embodiments, a sample can be deposited on one or both of the plates. In certain embodiments, in the closed configuration, at least a portion of the sample is squeezed into a layer by the two plates, where the average sample thickness is 200 μm or less.

[0894] In some embodiments, the QMAX card 100 includes a hinge 103 that connects the first plate 10 and the second plate 20 such that the two plates can pivot relative to each other. In some embodiments, the QMAX card includes a notch 105 that facilitates switching between the open and closed configurations. In some embodiments, one or both of the plates are transparent. In some embodiments, one or both of the plates are flexible. In some embodiments, the QMAX card 100 includes a heating / cooling layer 190. In certain embodiments, the heating / cooling layer 190 is configured to absorb electromagnetic waves and convert the energy to increase the temperature of the sample.

[0895] Figures 4A and 4B show a perspective view and a cross-sectional view of an embodiment of the device of the present invention. Figure 4A shows the device (also referred to as the "sample holder" of the system) 100 in the open configuration. As shown in Figure 4A, the sample holder 100 includes a first plate 10, a second plate 20, and a spacer mechanism (not shown). The first plate 10 and the second plate 20 each include an inner surface (11 and 21, respectively) and an outer surface (12 and 22, respectively). Each inner surface has a sample contact area (not shown) for contacting the fluid sample that the device is to process and / or analyze.

[0896] The first plate 10 and the second plate 20 can be moved relative to each other into different configurations. One such configuration is the open configuration, in which, as shown in Figure 4A, the first plate 10 and the second plate 20 are partially or fully separated, and the spacing between the first plate 10 and the second plate 20 (i.e., the distance between the inner surface 11 of the first plate and the inner surface 21 of the second plate) is not adjusted by the spacer mechanism. The open configuration allows a sample to be deposited on the first plate, the second plate, or both in the sample contact area.

[0897] As shown in FIG. 4A, the second plate 20 further includes a heating / cooling layer 112 in the sample contact area. The first plate 10 may also optionally or additionally include a heating / cooling layer 112. In some embodiments, the heating / cooling layer 112 is configured to effectively absorb radiation (such as electromagnetic waves) incident thereon. The absorption rate is 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 95% or more, 99% or more, 100% or less, 85% or less, 75% or less, 65% or less, or 55% or less, or within a range between any two values. The heating / cooling layer 112 is also configured to convert at least a majority of the absorbed radiation energy into heat (thermal energy). For example, the heating / cooling layer 112 is configured to release radiation in the form of heat after absorbing energy from electromagnetic waves. As used herein, the term “majority” or “substantially” refers to a percentage of 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 95% or more, 99% or more, 99% or more, or 99.9% or more.

[0898] FIGS. 3A and 3B show a sample card in a closed configuration, where the heating / cooling layer includes a heating zone directly heated / awaiting heating by a heating source; FIG. 3A shows a perspective view, while FIG. 3B shows a cross-sectional view. In some embodiments, the heating / cooling layer includes a heating zone being directly heated by a heating source. In some embodiments, the heating source emits electromagnetic radiation (waves), which reaches the heating / cooling layer with or without being modulated by a lens or other modulator. The area directly receiving such radiation (waves) is called the heating zone.

[0899] In some embodiments, the heating zone is smaller than the entire area of the heating / cooling layer. In some embodiments, the heating zone occupies about 1 / 1000, 1 / 500, 1 / 200, 1 / 100, 1 / 50, 1 / 20, 1 / 10, 1 / 5, 1 / 2, or 2 / 3 of the area of the heating / cooling layer, or within a range between any two values. In some embodiments, when a sample is loaded and pressed into a thin layer by two plates, the sample volume directly in the electromagnetic wave path or directly in contact with the area of the heating zone is called the heating volume. In some embodiments, due to the thinness of the sample layer and / or due to the excellent absorption performance of the heating / cooling layer, the sample in the heating volume can be quickly heated to the desired temperature. In some embodiments, the sample in the heating volume can also be quickly cooled to the desired temperature.

[0900] Biochemistry and Assay

[0901] The thermal cycler system and related methods of the present invention are used to facilitate chemical, biological, or medical assays or reactions. In some embodiments, the reaction requires a temperature change. In some embodiments, the reaction requires or preferably a rapid temperature change to avoid non-specific reactions and / or reduce waiting time. In certain embodiments, the systems and methods of the present invention are used to facilitate reactions that require cyclic temperature changes to amplify nucleotides in a fluid sample; these reactions include, but are not limited to, polymerase chain reaction (PCR). The following description uses PCR as an example to demonstrate the performance and uses of the thermal cycler system and methods of the present invention. However, it should be noted that some embodiments of the devices, systems, and methods described herein are also applicable to other assays and / or reactions that require temperature control and change.

[0902] In some embodiments, the assay (e.g., PCR) can be performed on an untreated sample. For example, the template for a PCR reaction can be provided by a sample obtained directly from a subject without additional processing. In some embodiments, the sample can be whole blood from an individual. In some embodiments, such a "one-step" method will allow for more convenient use of the devices described herein.

[0903] In some embodiments, sample 90 is a premixed reaction medium for polymerase chain reaction (PCR). For example, in certain embodiments, the reaction medium includes components such as, but not limited to: DNA template, two primers, DNA polymerase (e.g., Taq polymerase), deoxynucleoside triphosphates (dNTP), divalent cations (e.g., Mg 2+ ), monovalent cations (e.g., K + ), and a buffer solution. The specific components, the concentration of each component, and the total volume vary according to the rational design of the reaction. In some embodiments, a PCR assay requires many changes / alterations in the sample temperature between the following steps: (i) an optional initialization step that requires heating the sample to 92 - 98 °C; (2) a denaturation step that requires heating the sample to 92 - 98 °C; (3) an annealing step that requires reducing the sample temperature to 50 - 65 °C; (4) an extension (or elongation) step that requires heating the sample to 75 - 80 °C; (5) repeating steps (2) - (4) approximately 20 - 40 times; and (6) completing the assay and reducing the sample temperature to ambient temperature (e.g., room temperature) or cooling to approximately 4 °C. The specific temperature and specific time period for each step vary and depend on many factors, including but not limited to the length of the target sequence, the length of the primers, the cation concentration, and / or the GC percentage.

[0904] The thermal cycler system of the present invention achieves rapid temperature changes for PCR assays. For example, referring to FIGS. (A) and (B) of FIG. 3 and FIG. (B) of FIG. 4, in some embodiments, a sample 90 (e.g., a premixed reaction medium) is added to one or both plates 10 and 20 in an open configuration, and the plates are switched to a closed configuration to press the sample 90 into a thin layer having a thickness 102, which thickness 102 is adjusted by a spacer mechanism (not shown); a heating source 202 projects electromagnetic waves 210 onto the first plate 10 (e.g., specifically onto the heating / cooling layer 112); the heating / cooling layer 112 is configured to absorb the electromagnetic waves 210 and convert at least a majority of the electromagnetic waves 210 into heat, which heat raises the temperature of the sample; removal of the electromagnetic waves 210 causes the temperature of the sample 90 to decrease.

[0905] In some embodiments, by projecting the electromagnetic waves 210 onto the heating / cooling layer 112 or increasing the intensity of the electromagnetic waves, the thermal cycler system provides rapid heating (raising the temperature) for any or all of the initialization step, denaturation step, and / or extension / elongation step; in some embodiments, by removing the electromagnetic waves projected by the heating source 202 or decreasing the intensity of the electromagnetic waves, cooling in the annealing step and / or final cooling step is rapidly achieved. In some embodiments, the increase in the electromagnetic waves 210 or the intensity of the electromagnetic waves 210 produces a temperature rise rate of at least 80°C / s, 70°C / s, 60°C / s, 50°C / s, 45°C / s, 40°C / s, 35°C / s, 30°C / s, 25°C / s, 20°C / s, 18°C / s, 16°C / s, 14°C / s, 12°C / s, 10°C / s, 9°C / s, 8°C / s, 7°C / s, 6°C / s, 5°C / s, 4°C / s, 3°C / s, or 2°C / s, or within a range between any two values. In certain embodiments, the average temperature rise rate in the PCR assay is 10°C / s or higher. In some embodiments, the removal of the electromagnetic waves 210 or the decrease in the intensity of the electromagnetic waves 210 results in a temperature decrease rate of at least 80°C / s, 70°C / s, 60°C / s, 50°C / s, 45°C / s, 40°C / s, 35°C / s, 30°C / s, 25°C / s, 20°C / s, 18°C / s, 16°C / s, 14°C / s, 12°C / s, 10°C / s, 9°C / s, 8°C / s, 7°C / s, 6°C / s, 5°C / s, 4°C / s, 3°C / s, or 2°C / s, or within a range between any two values. In certain embodiments, the average temperature decrease rate in the PCR assay is 5°C / s or higher. As used herein, the term "temperature ramp rate" refers to the rate of temperature change between two preset temperatures. In some embodiments, the average rising or falling temperature for each step is different.

[0906] During the PCR process, after reaching the target temperature at any step, the sample needs to be maintained at the target temperature for a period of time. The thermal cycler system of the present invention provides a temperature holding function in the following ways: (1) adjusting the intensity of the electromagnetic wave 210, reducing the intensity of the electromagnetic wave 210 if the temperature has risen to the target temperature, or increasing the intensity of the electromagnetic wave 210 if the temperature has dropped to the target temperature, and / or (2) maintaining the target temperature by balancing the heat supplied to the sample and the heat removed from the sample.

[0907] FIG. 9 shows a cross-sectional view of an exemplary process of nucleic acid amplification using a device according to some embodiments. Examples of steps include (A) introducing a sample containing nucleic acid into the inner side of a first plate (substrate); (B) pressing a second plate (QMAX card) onto the inner surface of the first plate to form a closed configuration of the device, where necessary reagents for nucleic acid amplification are dried on the inner surface of the second plate; (C) accumulating nucleic acid amplification products in the chamber enclosed by the first and second plates.

[0908] When necessary, the sample can be introduced onto the first plate or the second plate, or even both. FIG. 9 herein provides an example of introducing the sample onto the inner surface of the first plate.

[0909] More specifically, in step (B), the second plate is pressed onto the inner surface of the first plate in contact with the sample to form a closed configuration of the device. The "second plate" may refer to a plate having periodic spacers on the inner surface in contact with the sample.

[0910] More specifically, in step (C), when the device is in the closed configuration, a heat source projects an electromagnetic wave onto a heating / cooling layer on the inner or outer surface of the first plate or the second plate or both. The heating / cooling layer is configured to absorb the electromagnetic wave and convert at least most of the energy from the electromagnetic wave into heat, and this heat is transferred to the sample in the enclosed chamber. In some embodiments, the heat source is programmed to adjust the temperature of the sample in the range from ambient temperature to 98°C. In some embodiments, for example, for conventional PCR, the sample is first heated to 98°C and then undergoes 15 to 40 repeated cycles at 94°C, 50 - 65°C, and 72°C. In some embodiments, for example, for isothermal amplification, the temperature of the sample is maintained at a constant temperature. In some embodiments, for example, when performing isothermal amplification via LAMP, the sample is heated to 60 - 65°C for about 1 - 70 minutes.

[0911] Figure 10 shows a cross-sectional view of an exemplary assay method for nucleic acid extraction and amplification using a card device according to some embodiments. Examples of the steps include (A) immobilizing capture probes on the inner surface of a first plate (substrate); (B) introducing a sample onto the inner surface of the first plate; (C) pressing a second plate onto the inner surface of the first plate to form a closed configuration of the device, wherein the necessary reagent 1 for facilitating the release and capture of nucleic acids is dried on the inner surface of the second plate; (D) capturing nucleic acids from the above sample on the inner surface of the first plate; (E) disassembling the second plate and cleaning the inner surface of the first plate with a sponge; (F) pressing a third plate onto the inner surface of the first plate, wherein the necessary reagent 2 for nucleic acid amplification is dried on the inner surface of the third plate; (G) accumulating nucleic acid amplification products in the chamber enclosed by the first plate and the third plate.

[0912] In some embodiments, in step (a), the capture probes are immobilized on the inner surface of the first plate. As used herein, a "capture probe" refers to an oligonucleotide having a length of 1-200 bp, preferably 5-50 bp, more preferably 10-20 bp. The capture probe may have a sequence complementary to the nucleic acid sequence relevant in the sample. In some embodiments, the same capture probes may be immobilized on the surface of the first plate. In some other embodiments, different capture probes having different base pair compositions are immobilized on the surface of the first plate. The capture probe may be DNA, or RNA, or both, but is preferably single-stranded DNA. As used herein, "immobilizing" refers to the process of anchoring the capture probe on the plate surface. In some embodiments, the capture probe is anchored by a covalent bond, wherein for example, the 5' or 3' end of the capture probe is modified to facilitate coating on the plate surface. Commonly used 3' end modifications include but are not limited to thiol, dithiol, amine, biotin, etc. In some other embodiments, the capture probe may be passively adsorbed on the substrate surface.

[0913] After immobilizing with the capture probe, the plate surface is blocked with a blocking agent solution. Suitable blocking agents include but are not limited to 6-mercapto-hexanol, bovine serum albumin, etc.

[0914] As shown in step (B) of Figure 10, a "sample" can be any nucleic acid with or without a sample, including but not limited to human body fluids such as whole blood, plasma, serum, urine, saliva, and sweat, as well as cell cultures (mammalian, plant, bacterial, fungal). The sample can be freshly obtained, or stored or processed in any desired or convenient manner, such as by dilution or addition of a buffer or other solution or solvent. Cellular structures such as human cells, animal cells, plant cells, bacterial cells, fungal cells, and virus particles may be present in the sample.

[0915] When necessary, the sample can be introduced onto the first plate or the second plate, or even both. Figure 10 herein provides an example of introducing the sample onto the inner surface of the first plate.

[0916] In some embodiments, in step (C), the second plate is pressed onto the inner surface of the first plate (substrate) in contact with the sample to form a closed configuration of the device. The necessary reagent 1 for nucleic acid amplification is in a dried form on the inner surface of the first plate or the second plate or both, or in a liquid form encapsulated, embedded or surrounded in a material (such as paraffin) that melts with increasing temperature.

[0917] In some embodiments, in step (D), after contact with the above-mentioned sample, the dried necessary reagent 1 is dissolved in the sample. The target nucleic acid released from the disrupted cell structure or present as cell-free nucleic acid or a combination thereof hybridizes with the complementary capture probe on the plate surface. The time for hybridization varies greatly depending on the specifications of the spacer on the inner surface of the plate. In some embodiments, for example, when using a plate with a spacer having a height of 30 μm, experimental data is indicated after 2 minutes, and the hybridization between the relevant nucleic acid and the immobilized capture probe reaches equilibrium. As used herein, "unhybridized nucleic acid" refers to nucleic acid not captured by the immobilized capture probe.

[0918] In some embodiments, in step (E) of FIG. 10, the second plate is separated from the first plate (substrate), and the surface of the first plate (substrate) is cleaned using a sponge. As used herein, "sponge" refers to a class of flexible porous materials that change pore size under different pressures. The sponge containing the washing buffer contacts the surface of the first plate to remove contaminants. In some embodiments, the sponge contacts the surface of the first plate once. In some other embodiments, the sponge contacts the surface of the first plate two or more times. As used herein, "contaminant" refers to a compound that is adverse to the nucleic acid amplification reaction, including but not limited to cell debris, proteins, non-specific nucleic acids, etc.

[0919] In some embodiments, in step (F) of FIG. 10, the third plate (QMAX card 2) is pressed onto the inner surface of the first plate in contact with the sample to form a closed configuration of the device. The necessary reagent 2 for nucleic acid amplification can be in a dried form on the inner surface of the first plate or the third plate or both, or in a liquid form encapsulated, embedded or surrounded in a material (such as paraffin) that melts with increasing temperature.

[0920] In some embodiments, in step (G) of FIG. 10, when the device is in the closed configuration, the heating source projects electromagnetic waves onto the heating / cooling layer on the inner or outer surface of the first plate or the third plate or both. The heating / cooling layer is configured to absorb the electromagnetic waves and convert at least most of the energy from the electromagnetic waves into heat, which is transferred to the sample in the enclosed chamber. In some embodiments, the heating source is programmed to adjust the temperature of the sample in the range from ambient temperature to 98 °C. In some embodiments, for example, for conventional PCR, the sample is first heated to 98 °C and then undergoes 15 - 40 repeated cycles at 94 °C, 50 - 65 °C, and 72 °C. In some embodiments, for example, for isothermal amplification, the temperature of the sample is maintained at a constant temperature. In some embodiments, for example, when isothermal amplification is performed by LAMP, the sample is heated to 60 - 65 °C for about 1 - 70 minutes.

[0921] In some embodiments, the sample contact area of one or both plates includes an extruded open flow monitoring surface structure (MSS), which is configured to monitor how much flow has occurred after COF. For example, in some embodiments, the MSS includes a shallow square array, which will cause friction to components in the sample (such as blood cells in blood). By examining the distribution of some components of the sample, information related to the flow of the sample and its components under COF can be obtained.

[0922] The depth of the MSS can be 1 / 1000, 1 / 100, 1 / 100, 1 / 5, 1 / 2 of the spacer height, or within the range between any two values, and is in a protruding shape or a hole shape.

[0923] Multiplexing

[0924] FIGS. 8A and 8B show perspective views of the sample holder 100 in the open configuration (FIG. 8A) and the closed configuration (FIG. 8B), where there are multiple sample contact areas on the plates, thus allowing the processing and analysis of multiple samples. As shown in FIGS. 8A and 8B, the thermal cycler system of the present invention includes a sample holder 100 and a thermal control unit 200; the sample holder 100 includes a first plate 10, a plurality of second plates 20, and a plurality of spacer mechanisms (not shown); the thermal control unit 200 includes a heating source 202 and a controller 204.

[0925] Referring to FIG. 8A, one or both of the plates (e.g., the second plate 20) include a plurality of sample contact areas (not labeled). In some embodiments, one or both of the plates (e.g., the second plate 20) include a plurality of heating / cooling layers 112. FIG. 8A shows the sample holder 100 in an open configuration, where the first plate 10 and the second plate 20 are partially or fully separated, allowing one or more samples to be deposited onto one or both of the plates. In the open configuration, the spacing between the first plate 10 and the second plate 20 is not adjusted by a spacing mechanism.

[0926] FIG. 8B shows the sample holder 100 in a closed configuration, where the inner surfaces of the two plates face each other, and the spacing 102 between the two plates is adjusted by a spacing mechanism (not shown). If one or more samples have been deposited on the plates, the plates are configured to press each sample into a layer, the thickness of which is adjusted by the spacing mechanism.

[0927] As shown in FIG. 8B, a plurality of first plates 10 are used to cover a portion of the second plate 20. For example, each first plate 10 covers a single sample contact area on which a sample is deposited. There is a spacing mechanism for each sample contact area, and the spacing mechanisms have different heights, resulting in different spacings 102 for each sample contact area and for different thicknesses of each sample layer. For example, the spacing mechanism is a columnar spacer; each sample contact area has a very uniform set of spacers; the heights of different sets of spacers are the same or different, resulting in the same or different sample layer thicknesses for different samples.

[0928] Referring to FIGS. 8A and 8B, in some embodiments, the controller 204 directs the heat source 202 to project electromagnetic waves 210 onto the second plate 20 (and thus onto the heating / cooling layer 112), where the electromagnetic waves 210 are absorbed by the heating / cooling layer 112 and converted into heat, causing a temperature change in the sample. In some embodiments, when there are multiple sample contact areas, multiple samples are processed and analyzed. For example, in certain embodiments, each sample is a pre-mixed PCR reaction medium with different components. One sample holder 100 is used to test different conditions for amplifying the same nucleotide and / or amplifying different nucleotides under the same or different conditions.

[0929] Other exemplary embodiments

[0930] AAA-1.1. A device for rapidly changing the temperature of a fluid sample, comprising:

[0931] A first plate (10), a second plate (20), a heating layer (112-1) and a cooling layer (112-2), wherein:

[0932] Each of the first plate and the second plate has a sample contact area on its respective inner surface for contacting a fluid sample; wherein the sample contact areas face each other, the average spacing between them is 200 μm or less, and are capable of contacting the sample and sandwiching the sample therebetween;

[0933] Heating layer:

[0934] Is positioned on the inner surface, outer surface or inside of one of the plates, and

[0935] Is configured to heat a relevant volume of the sample, wherein the relevant volume of the sample is part or all of the sample being heated to a desired temperature; and

[0936] Cooling layer:

[0937] Is located on the inner surface, outer surface or inside of one of the plates;

[0938] Is configured to cool a relevant sample volume; and

[0939] Contains a material layer with a thermal conductivity to heat capacity ratio of 0.6 cm 2 / sec or greater;

[0940] Wherein the distance between the cooling layer and the surface of the relevant sample volume is zero or less than the distance configured such that the thermal conductivity per unit area between the cooling layer and the surface of the relevant sample volume is equal to 70 W / (m 2 ·K) or greater; and

[0941] Wherein, in some embodiments, the heating layer and the cooling layer are the same material layer with a heating zone and a cooling zone, and wherein the heating zone and the cooling zone may have the same area or different areas.

[0942] AAA-1.2. A device for rapidly changing the temperature of a fluid sample, comprising:

[0943] A first plate (10), a second plate (20), a heating layer (112-1) and a cooling layer (112-2), wherein:

[0944] Each of the first plate and the second plate has a sample contact area on its respective inner surface for contacting a fluid sample; wherein the sample contact areas face each other, are separated by an average separation distance of 200 μm or less, and are capable of contacting the sample and sandwiching the sample therebetween;

[0945] Heating layer:

[0946] Is positioned on the inner surface, outer surface or inside of one of the plates,

[0947] configured to heat a relevant volume of a sample, where the relevant volume of the sample is part or all of the sample being heated to a desired temperature; and

[0948] Cooling layer:

[0949] Located on the inner surface, outer surface or inside of one of the plates;

[0950] configured to cool the relevant sample volume; and

[0951] comprising a layer of material having a thermal conductivity to heat capacity ratio of 0.6 cm 2 / sec or greater, where the layer with the high thermal conductivity to heat capacity ratio has an area greater than the lateral area of the sample volume;

[0952] where the distance between the cooling layer and the surface of the relevant sample volume is zero or less than the distance configured such that the thermal conductivity per unit area between the cooling layer and the surface of the relevant sample volume is equal to 70 W / (m 2 ·K) or greater; and

[0953] where, in some embodiments, the heating layer and the cooling layer are the same layer of material having a heating zone and a cooling zone, and where the heating zone and the cooling zone may have the same area or different areas.

[0954] AAA-1.3. Apparatus for rapidly changing the temperature of a fluid sample, comprising:

[0955] A first plate (10), a second plate (20), a heating layer (112-1) and a cooling layer (112-2), where:

[0956] The first plate and the second plate are movable relative to each other into different configurations;

[0957] Each of the first plate and the second plate has a sample contact area on its respective inner surface for contacting the fluid sample; where the sample contact areas face each other, are separated by an average spacing of 200 μm or less, and are capable of clamping the sample therebetween;

[0958] Heating layer:

[0959] Positioned on the inner surface, outer surface or inside of one of the plates,

[0960] configured to heat a relevant volume of a sample, where the relevant volume of the sample is part or all of the sample being heated to a desired temperature; and

[0961] Cooling layer:

[0962] Located on the inner surface, outer surface or inside of one of the plates;

[0963] configured to cool a related sample volume; and

[0964] comprising a material layer having a thermal conductivity to heat capacity ratio of 0.6 cm 2 / sec or greater;

[0965] wherein the distance between the cooling layer and the surface of the related sample volume is zero or less than a distance configured such that the thermal conductivity per unit area between the cooling layer and the surface of the related sample volume is equal to 70 W / (m 2 ·K) or greater;

[0966] wherein one of the configurations is an open configuration, wherein: the two plates are partially or fully separated and the average spacing between the plates is at least 300 μm;

[0967] wherein another of the configurations is a closed configuration, configured after depositing a fluid sample on one or both of the sample contact regions in the open configuration; and in the closed configuration: at least a portion of the sample is defined as a layer by the two plates, wherein the average sample thickness is 200 μm or less; and

[0968] wherein, in some embodiments, the heating layer and the cooling layer are the same material layer having a heating zone and a cooling zone, and wherein the heating zone and the cooling zone may have the same area or different areas.

[0969] AAA-1.4. A device for rapidly changing the temperature of a fluid sample, comprising:

[0970] a first plate (10), a second plate (20), spacers, a heating layer (112-1) and a cooling layer (112-2), wherein:

[0971] the first plate and the second plate are movable relative to each other into different configurations;

[0972] each of the first plate and the second plate has a sample contact region on its respective inner surface for contacting the fluid sample; wherein the sample contact regions face each other, the average separation distance between them is 200 μm or less, and are capable of contacting the sample and clamping the sample therebetween;

[0973] one or both of the plates comprise spacers, and the spacers are fixed to the inner surface of the corresponding plate;

[0974] the spacers have a predetermined substantially uniform height equal to or less than 200 microns, and a predetermined spacer spacing;

[0975] the heating layer:

[0976] is positioned on the inner surface, outer surface or inside of one of the plates,

[0977] configured to heat a relevant volume of a sample, wherein the relevant volume of the sample is part or all of the sample being heated to a desired temperature; and

[0978] Cooling layer:

[0979] Located on the inner surface, outer surface or inside of one of the plates;

[0980] configured to cool the relevant sample volume; and

[0981] comprising a material layer with a ratio of thermal conductivity to heat capacity of 0.6 cm 2 / sec or greater;

[0982] wherein the distance between the cooling layer and the surface of the relevant sample volume is zero or less than the distance configured such that the thermal conductivity per unit area between the cooling layer and the surface of the relevant sample volume is equal to 70 W / (m 2 ·K) or greater;

[0983] wherein one of the configuratio...

Claims

1. An apparatus allowing rapid thermal cycling, comprising: A first plate, said first plate comprising a polymer or glass material and having a thickness less than or equal to 100 µm; A second plate, said second plate comprising a polymer or glass material and having a thickness less than or equal to 100 µm, and each plate having a sample contact area for contacting a sample; One or both of the first plate and the second plate are flexible; the first plate and the second plate are movable relative to each other into different configurations; Spacers; And A heating / cooling layer, the heating / cooling layer is disposed on the sample contact area on the first plate or the second plate; the heating / cooling layer has a thermal conductivity, the thermal conductivity multiplied by the thickness of the heating / cooling layer is between 6×10 -5 W / K to 1.5×10 -4 Between W / K, Wherein the heating / cooling layer has a thickness of 15 µm or less and has a surface thermal radiation ability that is at least 50% of the surface thermal radiation ability of a black body, One of the configurations is an open configuration, in which: the two plates are partially or fully separated, the spacing between the plates is not adjusted by the spacers, and A fluid sample is deposited on one or both plates; Wherein, another configuration is a closed configuration, configured after depositing the fluid sample in the open configuration; and in the closed configuration: at least a portion of the fluid sample is squeezed by the two plates into a layer of uniform thickness, wherein the uniform thickness of the layer is defined by the sample contact surfaces of the plates and is adjusted by the plates and the spacers; The spacer is used to adjust the spacing between the first plate and the second plate, where at least one spacer is located within the sample contact area, and the ratio ISD of the fourth power of the spacer spacing ISD to the product of the thickness h and Young's modulus E of the flexible plate 4 / (hE) is less than or equal to 5×10 6 µm³ / GPa.

2. The apparatus according to claim 1, further comprising: A clamp, said clamp squeezing the first plate and the second plate to fix the two plates together, wherein squeezing applies pressure to the plates; and Wherein the pressure acts on the peripheral region of the area of the sample being clamped, thus reducing the sample outflow area.

3. A method for rapidly changing the temperature of a sample, comprising: i. Providing the apparatus of claim 1; ii. Depositing a fluid sample on one or both sample contact areas of the first plate and the second plate; iii. Pressing the plates by hand to clamp the sample between them and pressing at least a portion of the sample into a thin layer: iv. Changing and / or maintaining the temperature of the volume of the thin layer in the apparatus.

4. A method for rapidly changing the temperature of a sample, comprising: i. Providing the apparatus of claim 2; ii. Depositing a fluid sample on one or both sample contact areas of the first plate and the second plate; iii. Pressing the plates so that the sample contact areas face each other, Wherein the first plate and the second plate face each other in a parallel arrangement and are separated by a distance of 150 µm or less.

5. The apparatus of claim 1, Wherein: The first plate and the second plate are movable into different configurations.

6. The apparatus according to claim 1, wherein The second plate has a thickness less than or equal to 100 µm, wherein the second plate is separated from the first plate in a parallel arrangement by a distance less than or equal to the thickness of the second plate; and Wherein the heating / cooling layer is configured to receive electromagnetic radiation such that at least a portion of the liquid sample sandwiched between the first plate and the second plate is heated at a rate of at least 30 °C / sec.

7. The apparatus according to claim 1, comprising: The second plate has a thickness less than or equal to 100 µm, wherein the second plate is spaced from the first plate in a parallel arrangement by a distance less than or equal to the thickness of the second plate; and wherein when the heating / cooling layer does not receive electromagnetic radiation generated by a light source, at least a portion of the liquid sample sandwiched between the first plate and the second plate is cooled at a rate of at least 30 °C / sec.

8. The apparatus according to claim 1, comprising: wherein the inner surface of the second plate is spaced from the inner surface of the first plate in a parallel arrangement by a distance less than or equal to the thickness of the second plate; the heating / cooling layer is disposed on the inner or outer surface of the second plate; and a dry reagent layer is on the inner surface of the first plate.

9. A photothermal regulation system, comprising: the apparatus according to claim 1; a support frame configured to support at least one first plate and second plate; a housing having a first opening configured to receive the apparatus and at least one other opening; a light source configured to emit electromagnetic radiation towards the heating / cooling layer, wherein the heating / cooling layer is configured to absorb at least a portion of the electromagnetic radiation such that at least a portion of the liquid sample sandwiched between the first plate and the second plate is heated at a rate of at least 30 °C / sec, and wherein when the heating / cooling layer does not receive electromagnetic radiation generated by the light source, at least a portion of the liquid sample sandwiched between the first plate and the second plate is cooled at a rate of at least 30 °C / sec, and wherein the system consumes less than 500 mW of power.

10. A photothermal regulation system, comprising: the apparatus according to claim 1, a support frame configured to support at least one first plate and second plate; and a light source configured to emit electromagnetic radiation towards the heating / cooling layer, wherein when the heating / cooling layer does not receive electromagnetic radiation generated by the light source, at least a portion of the liquid sample sandwiched between the first plate and the second plate is cooled at a rate of at least 30 °C / sec.

11. A photothermal regulation system, comprising: the apparatus according to claim 1, a light source configured to emit electromagnetic radiation towards the heating / cooling layer, wherein the system consumes less than 500 mW of power.

12. A photothermal regulation system, comprising: the apparatus according to claim 1, a support frame configured to support at least one of the first plate and the second plate; a housing having a first opening configured to receive the apparatus and at least one other opening; and a light source configured to emit electromagnetic radiation through the at least one other opening of the housing and towards the heating / cooling layer, wherein when the heating / cooling layer does not receive electromagnetic radiation generated by the light source, the liquid sample sandwiched between the first plate and the second plate is cooled at a rate of at least 30 °C / sec.

13. A method of using the apparatus according to claim 1, comprising: Place a second plate on the first plate such that a fluid sample is sandwiched between the first and second plates at a thickness determined by one or more spacers located on at least one of the first and second plates; Activate a heat source configured to radiate electromagnetic radiation towards a heating layer located on the first or second plate; and Use at least the heating layer to heat at least a portion of the fluid sample at a rate of at least 30 °C / sec.

14. A method of using the apparatus of claim 1, comprising: Place a second plate on the first plate such that a fluid sample is sandwiched between the first and second plates at a thickness determined by one or more spacers located on at least one of the first and second plates; Activate a heat source configured to radiate electromagnetic radiation towards a heating / cooling layer located on the first or second plate for a given period of time; Deactivate the heat source after the given period of time, wherein at least a portion of the fluid sample cools at a rate of at least 30 °C / sec after deactivation.

15. A method of using the apparatus of claim 1, comprising: Place a second plate on the first plate such that a fluid sample is sandwiched between the first and second plates at a thickness determined by one or more spacers located on at least one of the first and second plates; Activate a heat source configured to radiate electromagnetic radiation towards a heating layer located on the first or second plate, wherein the heat source consumes less than 500 mW of power; and Use at least the heating layer to heat at least a portion of the fluid sample.

16. A method of amplifying nucleic acid, comprising: Obtain the apparatus of claim 1; Deposit a fluid sample containing nucleic acid on the first plate of the apparatus; Place a second plate on the first plate such that the fluid sample is sandwiched between the first and second plates, wherein a reagent for nucleic acid amplification is present on the inner surface of the second plate; Activate a heat source configured to radiate electromagnetic radiation towards a heating layer located on the first or second plate; Use at least the heating layer to heat at least a portion of the fluid sample at a rate of at least 30 °C / sec; and Accumulate nucleic acid amplification products in at least a portion of the fluid sample sandwiched between the first and second plates.

17. A method of amplifying nucleic acid, comprising: Obtain the apparatus of claim 1, Deposit a fluid sample containing nucleic acid on the first plate of the apparatus; Place a second plate on the first plate such that the fluid sample is sandwiched between the first and second plates, wherein a reagent for nucleic acid amplification is present on the inner surface of the second plate; Amplify the nucleic acid in the sample by performing one or more PCR cycles, wherein each PCR cycle comprises a denaturation step, an annealing step, and an extension step; wherein one or more of the denaturation step, the annealing step, and / or the extension step comprises: Activate a heat source configured to radiate electromagnetic radiation towards a heating layer located on the first or second plate; and Use at least the heating layer to heat at least a portion of the fluid sample at a rate of at least 30 °C / sec.

18. A method for amplifying nucleic acid, comprising: obtaining the device according to claim 1; depositing a fluid sample containing nucleic acid on the first plate of the device; placing a second plate on the first plate such that the fluid sample is sandwiched between the first plate and the second plate, wherein a reagent for nucleic acid amplification is present on the inner surface of the second plate; activating a heat source within a given time period, the heat source being configured to radiate electromagnetic radiation towards a heating / cooling layer located on the first plate or the second plate; deactivating the heat source after the given time period, wherein at least a portion of the fluid sample adjacent to the heating / cooling layer cools at a rate of at least 30 °C / sec after deactivation; and accumulating nucleic acid amplification products in at least a portion of the fluid sample sandwiched between the first plate and the second plate.

19. A method for amplifying nucleic acid, comprising: obtaining the device according to claim 1; depositing a fluid sample containing nucleic acid on the first plate of the device; placing a second plate on the first plate such that the fluid sample is sandwiched between the first plate and the second plate, wherein a reagent for nucleic acid amplification is present on the inner surface of the second plate; amplifying the nucleic acid in the sample by performing one or more PCR cycles, wherein each PCR cycle comprises a denaturation step, an annealing step, and an extension step; wherein one or more of the denaturation step, the annealing step, and / or the extension step comprises: activating a heat source, the heat source being configured to radiate electromagnetic radiation towards a heating layer located on the first plate or the second plate; and deactivating the heat source after a given time period, wherein at least a portion of the fluid sample adjacent to the heating / cooling layer cools at a rate of at least 30 °C / sec after deactivation; and accumulating nucleic acid amplification products in at least a portion of the fluid sample sandwiched between the first plate and the second plate.

20. A method for amplifying nucleic acid, comprising: obtaining the device according to claim 1; depositing a fluid sample containing nucleic acid on the first plate of the device; placing a second plate on the first plate such that the fluid sample is sandwiched between the first plate and the second plate with a thickness determined by one or more spacers located on at least one of the first plate and the second plate, wherein a reagent for nucleic acid amplification is present on the inner surface of the second plate; activating a heat source, the heat source being configured to radiate electromagnetic radiation towards a heating layer located on the first plate or the second plate, wherein the heat source consumes a power of less than 500 mW; heating at least a portion of the fluid sample using at least the heating layer; and accumulating nucleic acid amplification products in at least a portion of the fluid sample sandwiched between the first plate and the second plate.

21. A method for amplifying nucleic acid, comprising: obtaining the device according to claim 1; depositing a fluid sample containing nucleic acid on the first plate of the device; placing a second plate on the first plate such that the fluid sample is sandwiched between the first plate and the second plate with a thickness determined by one or more spacers located on at least one of the first plate and the second plate, wherein a reagent for nucleic acid amplification is present on the inner surface of the second plate; Amplify nucleic acids in a sample by performing one or more PCR cycles, where each PCR cycle includes a denaturation step, an annealing step, and an extension step; where one or more of the denaturation step, the annealing step, and / or the extension step include: Activating a heat source configured to radiate electromagnetic radiation to a heating layer located on the first plate or the second plate, where the heat source consumes less than 500 mW of power; Using at least the heating layer to heat at least a portion of the fluid sample; and Accumulating nucleic acid amplification products in at least a portion of the fluid sample sandwiched between the first plate and the second plate.

22. A method for detecting the presence or absence of a target nucleic acid sequence in a sample, comprising: Obtaining the device of claim 1, Depositing a fluid sample containing nucleic acids on the first plate of the device; Placing a second plate on the first plate such that the fluid sample is sandwiched between the first plate and the second plate, where reagents for nucleic acid amplification are present on the inner surface of the second plate, and where the reagents include primers capable of hybridizing to the target nucleic acid; Activating a heat source configured to radiate electromagnetic radiation to a heating layer located on the first plate or the second plate; Using at least the heating layer to heat at least a portion of the fluid sample at a rate of at least 30 °C / sec; and detecting whether the fluid sample contains amplification products of the target nucleic acid sequence.

23. A method for detecting the presence or absence of a target nucleic acid sequence in a sample, comprising: Obtaining the device of claim 1, Depositing a fluid sample containing nucleic acids on the first plate of the device; Placing a second plate on the first plate such that the fluid sample is sandwiched between the first plate and the second plate, where reagents for nucleic acid amplification are present on the inner surface of the second plate, and where the reagents include primers capable of hybridizing to the target nucleic acid; Activating a heat source for a given period of time, the heat source being configured to radiate electromagnetic radiation to a heating / cooling layer located on the first plate or the second plate; Deactivating the heat source after the given period of time, where at least a portion of the fluid sample adjacent to the heating / cooling layer cools at a rate of at least 30 °C / sec after deactivation; and Detecting whether the fluid sample contains amplification products of the target nucleic acid sequence.

24. A method for detecting the presence or absence of a target nucleic acid sequence in a sample, comprising: Obtaining the device of claim 1, Depositing a fluid sample containing nucleic acids on the first plate of the device; Placing a second plate on the first plate such that the fluid sample is sandwiched between the first plate and the second plate, where reagents for nucleic acid amplification are present on the inner surface of the second plate, and where the reagents include primers capable of hybridizing to the target nucleic acid; Activating a heat source configured to radiate electromagnetic radiation to a heating layer located on the first plate or the second plate, where the heat source consumes less than 500 mW of power; Using at least the heating layer to heat at least a portion of the fluid sample; and Detecting whether the fluid sample contains amplification products of the target nucleic acid sequence.

25. A method for detecting the presence or absence of an analyte in a sample, comprising: Obtaining the device of claim 1, Deposit a fluid sample on the first plate of the device; Place a second plate on the first plate such that the fluid sample is sandwiched between the first plate and the second plate, wherein a reagent for detecting an analyte is present on the inner surface of the second plate; Activate a heat source configured to radiate electromagnetic radiation towards a heating layer located on the first plate or the second plate; Use at least the heating layer to heat at least a portion of the fluid sample at a rate of at least 30 °C / sec; And detect whether the fluid sample contains the analyte.

26. A method for detecting the presence or absence of an analyte in a sample, comprising: Obtain the device of claim 1, Deposit a fluid sample contained thereon on the first plate of the device; Place a second plate on the first plate such that the fluid sample is sandwiched between the first plate and the second plate, wherein a reagent for detecting an analyte is present on the inner surface of the second plate; Activate a heat source configured to radiate electromagnetic radiation towards a heating / cooling layer located on the first plate or the second plate for a given period of time; Deactivate the heat source after the given period of time, wherein at least a portion of the fluid sample adjacent to the heating / cooling layer cools at a rate of at least 30 °C / sec after deactivation; And Detect whether the fluid sample contains the analyte.

27. A method for detecting the presence or absence of an analyte in a sample, comprising: Obtain the device of claim 1, Deposit a fluid sample on the first plate of the device; Place a second plate on the first plate such that the fluid sample is sandwiched between the first plate and the second plate, wherein a reagent for detecting an analyte is present on the inner surface of the second plate; Activate a heat source configured to radiate electromagnetic radiation towards a heating layer located on the first plate or the second plate, wherein the heat source consumes less than 500 mW of power; Use at least the heating layer to heat at least a portion of the fluid sample; And Detect whether the fluid sample contains the analyte.

28. The method according to claim 3, wherein during the thermal cycling, the proportion of the radiative cooling of the heating / cooling layer to the total cooling of the sample and the sample holder is at least 30% to 99%.

29. The device according to claim 1, Characterized in that: The two plates are movable relative to each other into different configurations One or both plates include height-uniform spacers; One configuration is an open configuration, wherein the two plates are partially or fully separated and the spacing between the plates is not adjusted by the spacers, thereby allowing a sample to be deposited on one or both plates; And Another configuration is a closed configuration configured such that at least a portion of the sample can be sandwiched between the first plate and the second plate to form a layer of uniform thickness, wherein the layer is bounded by the inner surfaces of the first plate and the second plate and is adjusted by the plates and the spacers.

30. The device according to claim 8, further comprising a light-absorbing layer provided on the heating / cooling layer, wherein the light-absorbing layer has an average light absorptivity of at least 30%.

31. The device according to claim 30, wherein the light-absorbing layer comprises a black coating.

32. The device according to claim 1, wherein the first plate is movable relative to the second plate to form different configurations including an open configuration and a closed configuration, wherein, in the open configuration, the first plate and the second plate are partially or completely separated, and the average spacing between the first plate and the second plate is at least 300 μm. The closed configuration is the configuration after depositing a sample onto one or two upper sample contact areas in the open configuration, and in the closed configuration: at least part of the sample is confined between the two plates to form a layer, and the thickness of the layer is 200 microns or less.

33. The device according to claim 32, wherein in the closed configuration, the spacing between the first plate and the second plate is 30 or less.

34. The device according to claim 32, wherein in the closed configuration, the spacing between the first plate and the second plate is 20 or less.

35. The device according to claim 32, wherein in the closed configuration, the spacing between the first plate and the second plate is 10 or less.

36. The device according to claim 1, wherein the thickness of the heating / cooling layer is less than or equal to 3 μm.

37. The device according to claim 1, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

38. The device according to claim 1, wherein the spacer is a plurality of spherical spacers disposed between the first plate and the second plate.

39. The device according to claim 1, wherein the spacer has a height of 10 μm, and the spacer is disposed between the first plate and the second plate.

40. The device according to claim 32, wherein the distance between the first plate and the second plate is less than or equal to 100 μm.

41. The device according to claim 1, further comprising a hinge configured to connect the first plate and the second plate and coupled to an edge of the first plate or the second plate.

42. The device according to claim 1, further comprising a light absorption layer disposed on the heating / cooling layer, wherein the light absorption layer has an average light absorption rate of at least 30%.

43. The device according to claim 42, wherein the light absorption layer comprises a black coating.

44. The method according to claim 13, wherein at least a portion of the sample is deposited along a path of electromagnetic radiation.

45. The device according to claim 1, wherein the spacers are arranged in a periodic array.

46. The device according to claim 1, wherein the height of the spacer is 100 μm, and the spacer is disposed between the first plate and the second plate.

47. The method according to claim 3, wherein at least a portion of the liquid sample comprises a volume of the sample adjacent to the heating / cooling layer.

48. The device according to claim 8, wherein the dried reagent layer comprises reagents for nucleic acid amplification.

49. The system according to claim 10, wherein the device further comprises a light absorption layer disposed on the heating / cooling layer, wherein the light absorption layer has an average light absorption rate of at least 30%.

50. The system according to claim 49, wherein the light absorption layer comprises a black coating.

51. The system according to claim 10, wherein the first plate is movable relative to the second plate.

52. The system according to claim 10, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

53. The system according to claim 10, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

54. The system according to claim 10, wherein the light source comprises a light-emitting diode.

55. The system according to claim 10, further comprising an optical waveguide configured to emit the electromagnetic radiation from the light source to the heating / cooling layer.

56. The system according to claim 9, wherein at least one other opening of the housing is configured to be aligned with at least a portion of the sample sandwiched between the first plate and the second plate when the device is placed in the housing via the first opening.

57. The system according to claim 10, wherein the support frame is configured to support at least the first plate or the second plate along the perimeter of the first plate or the second plate.

58. The system according to claim 9, wherein the first plate is movable relative to the second plate.

59. The system according to claim 9, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

60. The system according to claim 9, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

61. The system according to claim 9, wherein the light source comprises a light-emitting diode.

62. The system according to claim 61, wherein, the light-emitting diode comprises a blue LED.

63. The system according to claim 9, further comprising an optical waveguide configured to emit the electromagnetic radiation from the light source to the heating / cooling layer.

64. The system according to claim 9, wherein the support frame is configured to support at least the first plate or the second plate along the perimeter of the first plate or the second plate.

65. The system according to claim 9, wherein the device further comprises a light absorption layer disposed on the heating / cooling layer, and wherein the light absorption layer has an average light absorption rate of at least 30%.

66. The system according to claim 65, wherein the light absorption layer comprises a black coating.

67. The system according to claim 11, wherein the light source comprises a light-emitting diode.

68. The system according to claim 67, the light-emitting diode comprises a blue LED.

69. The system according to claim 11, further comprising an optical waveguide configured to emit the electromagnetic radiation from the light source to the heating / cooling layer.

70. The system according to claim 11, further comprising a support frame configured to: support at least the first plate or the second plate along the perimeter of the first plate or the second plate.

71. The system according to claim 11, wherein the device further comprises a light absorption layer disposed on the heating / cooling layer, and wherein the light absorption layer has an average light absorption rate of at least 30%.

72. The system according to claim 71, wherein the light absorption layer comprises a black coating.

73. The system according to claim 11, wherein the first plate is movable relative to the second plate.

74. The system according to claim 11, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

75. The system according to claim 11, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

76. The system according to claim 12, wherein the light source comprises a light-emitting diode.

77. The system according to claim 76, wherein, the light-emitting diode comprises a blue LED.

78. The system according to claim 12, further comprising an optical waveguide configured to guide the electromagnetic radiation from the light source to the heating / cooling layer.

79. The system according to claim 12, wherein the support frame is configured to support at least the first plate or the second plate along the perimeter of the first plate or the second plate.

80. The method according to claim 3, wherein the first plate or the second plate further comprises a light-absorbing layer disposed on the heating layer, and wherein the light-absorbing layer has an average light absorption rate of at least 30%.

81. The method according to claim 80, wherein the light-absorbing layer comprises a black coating.

82. The method according to claim 3, further comprising closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

83. The method according to claim 3, wherein the thickness of the heating layer is less than or equal to 3 µm.

84. The method according to claim 3, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

85. The method according to claim 3, wherein activating the heat source comprises activating an LED to radiate light onto the heating layer.

86. The method according to claim 3, further comprising controlling the output of the LED based on the measured or estimated temperature of a portion of the sample.

87. The method according to claim 3, further comprising supporting the perimeter of the first plate or the second plate on a support frame.

88. The method according to claim 4, wherein the first plate or the second plate further comprises a light-absorbing layer disposed on the heating / cooling layer, and wherein the light-absorbing layer has an average light absorption rate of at least 30%.

89. The method according to claim 88, wherein the light-absorbing layer comprises a black coating.

90. The method according to claim 4, further comprising closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

91. The method according to claim 4, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

92. The method according to claim 4, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

93. The method according to claim 4, wherein activating the heat source comprises: activating an LED to radiate light onto the heating / cooling layer.

94. The method according to claim 4, further comprising controlling the output of the LED based on the measured or estimated temperature of a portion of the fluid sample.

95. The method according to claim 4, further comprising supporting the perimeter of the first plate or the second plate on a support frame.

96. The method according to claim 16, wherein the first plate or the second plate further comprises a light-absorbing layer disposed on the heating layer, and wherein the light-absorbing layer has an average light absorption rate of at least 30%.

97. The method according to claim 96, wherein the light-absorbing layer comprises a black coating.

98. The method according to claim 16, further comprising closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

99. The method according to claim 16, wherein the thickness of the heating layer is less than or equal to 3 µm.

100. The method according to claim 16, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

101. The method according to claim 16, wherein the activation heat source comprises activating an LED to radiate light towards the heating layer.

102. The method according to claim 16, further comprising controlling the output of the LED based on the measured or estimated temperature of a portion of the fluid sample.

103. The method according to claim 16, further comprising using a beam expander to expand the electromagnetic radiation before the electromagnetic radiation reaches the heating layer.

104. The method according to claim 16, further comprising supporting the perimeter of the first plate or the second plate on a support frame.

105. The method according to claim 17, wherein the first plate or the second plate further comprises a light absorption layer disposed on the heating / cooling layer, wherein the light absorption layer comprises a black coating and has an average light absorption rate of at least 30%.

106. The method according to claim 17, further comprising closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

107. The method according to claim 17, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

108. The method according to claim 17, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

109. The method according to claim 17, wherein the activation heat source comprises: activating an LED to radiate light towards the heating / cooling layer.

110. The method according to claim 17, further comprising controlling the output of the LED based on the measured or estimated temperature of a portion of the fluid sample.

111. The method according to claim 17, further comprising using a beam expander to expand the electromagnetic radiation before the electromagnetic radiation reaches the heating layer.

112. The method according to claim 17, further comprising supporting the perimeter of the first plate or the second plate on a support frame.

113. The method according to claim 18, wherein the first plate or the second plate further comprises a light absorption layer disposed on the heating / cooling layer, wherein the light absorption layer has an average light absorption rate of at least 30%.

114. The method according to claim 113, wherein the light absorption layer comprises a black coating.

115. The method according to claim 18, further comprising closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

116. The method according to claim 18, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

117. The method according to claim 18, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

118. The method according to claim 18, wherein the activation heat source comprises: activating an LED to radiate light towards the heating / cooling layer.

119. The method according to claim 18, further comprising controlling the output of the LED based on the measured or estimated temperature of a portion of the fluid sample.

120. The method according to claim 18, further comprising using a beam expander to expand the electromagnetic radiation before the electromagnetic radiation reaches the heating layer.

121. The method according to claim 18, further comprising supporting the perimeter of the first plate or the second plate on a support frame.

122. The method according to claim 121, wherein the light absorption layer comprises a black coating.

123. The method according to claim 19, further comprising closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

124. The method according to claim 19, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

125. The method according to claim 19, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

126. The method according to claim 19, wherein activating the heat source comprises: activating an LED to radiate light towards the heating / cooling layer.

127. The method according to claim 19, further comprising controlling the output of the LED based on a measured or estimated temperature of a portion of the fluid sample.

128. The method according to claim 19, further comprising using a beam expander to expand the electromagnetic radiation before the electromagnetic radiation reaches the heating layer.

129. The method according to claim 19, further comprising supporting the periphery of the first plate or the second plate on a support frame.

130. The method according to claim 20, wherein the first plate or the second plate further comprises a light absorption layer disposed on the heating / cooling layer, and wherein the light absorption layer has an average light absorption rate of at least 30%.

131. The method according to claim 130, wherein the light absorption layer comprises a black coating.

132. The method according to claim 20, further comprising closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

133. The method according to claim 20, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

134. The method according to claim 20, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

135. The method according to claim 20, wherein activating the heat source comprises: activating an LED to radiate light towards the heating / cooling layer.

136. The method according to claim 20, further comprising controlling the output of the LED based on a measured or estimated temperature of a portion of the fluid sample.

137. The method according to claim 20, further comprising using a beam expander to expand the electromagnetic radiation before the electromagnetic radiation reaches the heating layer.

138. The method according to claim 20, further comprising supporting the periphery of the first plate or the second plate on a support frame.

139. The method according to claim 21, wherein the first plate or the second plate further comprises a light absorption layer disposed on the heating / cooling layer, and wherein the light absorption layer has an average light absorption rate of at least 30%.

140. The method according to claim 139, wherein the light absorption layer comprises a black coating.

141. The method according to claim 21, further comprising closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

142. The method according to claim 21, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

143. The method according to claim 21, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

144. The method according to claim 21, wherein activating the heat source comprises: activating an LED to radiate light towards the heating / cooling layer.

145. The method according to claim 21 further comprises controlling the output of the LED based on the measured or estimated temperature of a portion of the fluid sample.

146. The method according to claim 21 further comprises using a beam expander to expand the electromagnetic radiation before the electromagnetic radiation reaches the heating layer.

147. The method according to claim 21 further comprises supporting the periphery of the first plate or the second plate on a support frame.

148. The method according to claim 22, wherein the first plate or the second plate further comprises a light absorption layer disposed on the heating / cooling layer, and the light absorption layer has an average light absorption rate of at least 30%.

149. The method according to claim 148, wherein the light absorption layer comprises a black coating.

150. The method according to claim 22 further comprises closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

151. The method according to claim 22, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

152. The method according to claim 22, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

153. The method according to claim 22, wherein activating the heat source comprises: activating the LED to irradiate light onto the heating / cooling layer.

154. The method according to claim 22 further comprises controlling the output of the LED based on the measured or estimated temperature of a portion of the fluid sample.

155. The method according to claim 22 further comprises using a beam expander to expand the electromagnetic radiation before the electromagnetic radiation reaches the heating layer.

156. The method according to claim 22 further comprises supporting the periphery of the first plate or the second plate on a support frame.

157. The method according to claim 23, wherein the first plate or the second plate further comprises a light absorption layer disposed on the heating / cooling layer, and the light absorption layer has an average light absorption rate of at least 30%.

158. The method according to claim 157, wherein the light absorption layer comprises a black coating.

159. The method according to claim 23 further comprises closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

160. The method according to claim 23, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

161. The method according to claim 23, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

162. The method according to claim 23, wherein activating the heat source comprises: activating the LED to irradiate light onto the heating / cooling layer.

163. The method according to claim 23 further comprises controlling the output of the LED based on the measured or estimated temperature of a portion of the fluid sample.

164. The method according to claim 23 further comprises using a beam expander to expand the electromagnetic radiation before the electromagnetic radiation reaches the heating layer.

165. The method according to claim 23 further comprises supporting the periphery of the first plate or the second plate on a support frame.

166. The method according to claim 24, wherein the first plate or the second plate further comprises a light absorption layer disposed on the heating / cooling layer, and the light absorption layer has an average light absorption rate of at least 30%.

167. The method according to claim 166, wherein the light absorption layer comprises a black coating.

168. The method according to claim 24, further comprising closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

169. The method according to claim 24, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

170. The method according to claim 24, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

171. The method according to claim 24, wherein activating the heat source comprises: activating an LED to radiate light onto the heating / cooling layer.

172. The method according to claim 24, further comprising controlling the output of the LED based on the measured or estimated temperature of a portion of the fluid sample.

173. The method according to claim 24, further comprising using a beam expander to expand the electromagnetic radiation before the electromagnetic radiation reaches the heating layer.

174. The method according to claim 24, further comprising supporting the periphery of the first plate or the second plate on a support frame.

175. The method according to claim 25, wherein the first plate or the second plate further comprises a light absorption layer disposed on the heating / cooling layer, and the light absorption layer has an average light absorption rate of at least 30%.

176. The method according to claim 175, wherein the light absorption layer comprises a black coating.

177. The method according to claim 25, further comprising closing the second plate on the first plate using a hinge connected between the first plate and the second plate.

178. The method according to claim 25, wherein the thickness of the heating / cooling layer is less than or equal to 3 µm.

179. The method according to claim 25, wherein the entire main surface area of at least one of the first plate and the second plate is 400 mm 2 .

180. The method according to claim 25, wherein activating the heat source comprises: activating an LED to radiate light onto the heating / cooling layer.

181. The method according to claim 25, further comprising controlling the output of the LED based on the measured or estimated temperature of a portion of the fluid sample.

182. The method according to claim 25, further comprising using a beam expander to expand the electromagnetic radiation before the electromagnetic radiation reaches the heating layer.

183. The method according to claim 25, further comprising supporting the periphery of the first plate or the second plate on a support frame.

184. A kit for allowing rapid thermal cycling, comprising: (i) the device according to claim 8; and (ii) a premixed polymerase chain reaction medium.

185. The kit according to claim 184, wherein the premixed polymerase chain reaction medium comprises: a DNA template, two primers, a DNA polymerase, deoxynucleoside triphosphates, divalent cations, monovalent cations, and a buffer solution.

186. A device for measuring a thin layer of a fluid sample, comprising: a first plate, a second plate, a spacer, and a clamp, wherein: i. The first plate and the second plate are movable relative to each other into different configurations, including an open configuration and a closed configuration; ii. Each plate includes a sample contact area on its respective surface for contacting a fluid sample; iii. One or both plates include spacers fixed to the respective plates; iv. The spacers have a predetermined uniform height equal to or less than 200 micrometers, wherein at least one of the spacers is within the sample contact area; and v. A heating layer is configured to heat the fluid sample; wherein the heating layer is (a) on or near an inner or outer surface of one plate; or within one plate, and (b) capable of being heated by a heat source, wherein the heat source transfers thermal energy to the heating layer by light, electricity, radio frequency radiation, or a combination thereof; wherein in the open configuration, the two plates are partially or fully separated, the spacing between the plates is not adjusted by the spacers, and the sample is deposited on one or both plates; wherein in the closed configuration configured after the sample is deposited in the open configuration, at least a portion of the sample is squeezed by the two plates into a layer of uniform thickness and is stagnant relative to the plates, wherein the uniform thickness of the layer is defined by the sample contact areas of the two plates and is adjusted by the plates and the spacers; wherein the first plate and the second plate are configured to confine at least a portion of the sample in a layer of very uniform thickness of 0.1 - 200 µm and stagnant relative to the plates; wherein the first plate has a thickness of 500 µm or less, and the second plate has a thickness of 5 mm or less; The spacer is used to adjust the distance between the first plate and the second plate, where at least one spacer is located within the sample contact area, and the ratio ISD of the fourth power of the spacer distance ISD to the product of the thickness h and Young's modulus E of the flexible plate 4 / (hE) is less than or equal to 5×10 6 µm³ / GPa.

187. The apparatus according to claim 186, wherein the plate and the sample thickness are configured to allow the temperature of the sample to change at a rate of 10 °C / s or higher.

188. The device according to claim 187, wherein the clamp presses the first plate and the second plate to fix the two plates together in the closed configuration, and the pressure of the clamp inserted on the plates is 0.01 kg / cm 2 or higher.

189. The apparatus according to claim 188, wherein the heating layer is on or near one of the plates, has an absorption coefficient of 60% or higher, and has a thickness of less than 2 mm.

190. The apparatus according to claim 189, further comprising a radiation absorption layer, the radiation absorption layer being located near at least a portion of the sample of uniform thickness, and the area of at least a portion of the sample and the radiation absorption layer being greater than the uniform thickness.

191. The apparatus according to claim 190, wherein the area of at least a portion of the sample and the radiation absorption layer is greater than the uniform thickness of the sample.

192. The apparatus according to claim 191, wherein the apparatus has a plate with a thickness of 100 µm or less.

193. A system for rapidly changing the temperature of a thin fluid sample layer, comprising: i. The apparatus according to claim 190, ii. A radiation source, wherein the radiation source is configured to radiate electromagnetic waves significantly absorbed by the radiation absorption layer; and iii. A controller configured to control the radiation source and change the temperature of the sample.

194. A method for rapidly changing the temperature of a thin fluid sample layer, comprising: i. Providing the apparatus according to claim 190, or the system according to claim 193; ii. Deposit the fluid sample on one or both plates of the device; iii. After ii, press the plates into a closed configuration, wherein the plates squeeze at least a portion of the sample into a thin layer with a thickness less than 200 µm; and iv. Alter and maintain the temperature of the sample layer by changing the presence, intensity, wavelength, frequency, and / or angle of the electromagnetic wave from the radiation source.

195. The device according to claim 188, wherein the clamp is configured to include a thermal insulator layer to reduce heat conduction between the clamp and the plate, and wherein the thermal insulator layer comprises a material with a thermal conductivity of 2 W / mK.

196. The device according to claim 188, wherein the clamp is configured to include a thermal insulator layer to reduce the thermal mass required to heat or cool the sample, and wherein the thermal insulator layer comprises a material with a thermal conductivity of 2 W / mK.

197. The method according to claim 194, wherein in the closed configuration, the clamp is configured to have thermally conductive contact with a portion of the surface of the plate.

198. The method according to claim 194, wherein in the closed configuration, the clamp only has thermally conductive contact with the peripheral surface area of the plate.

199. The method according to claim 194, wherein in the closed configuration, the clamp only has thermally conductive contact with the surface area of the plate, and wherein the surface area is outside the portion of the sample where the nucleic acid to be amplified is located.

200. The device according to claim 188, wherein the clamp includes a window of transparency, and the window allows external light to shine on the plate or allows light inside the plate to come out, and wherein the transparency refers to higher than 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or within the range between any two of these values.

201. The device according to claim 188, wherein the clamp applies pressure to squeeze the first plate and the second plate, and the pressure is 0.01 kg / cm², 0.1 kg / cm², 0.5 kg / cm², 1 kg / cm², 2 kg / cm², kg / cm², 5 kg / cm², 10 kg / cm², 20 kg / cm², 30 kg / cm², 40 kg / cm², 50 kg / cm², 60 kg / cm², 100 kg / cm², 150 kg / cm², 200 kg / cm², 400 kg / cm², or within the range between any two of these values.

202. The device according to claim 186, further comprising a clamp that squeezes the first plate and the second plate together in the closed configuration, and further comprising a sealing material located between at least a portion of the first plate and the second plate, and the pressure of the clamp inserted on the plate is 0.01 kg / cm² or higher.

203. The method according to claim 194, wherein the change in the temperature of the sample is a thermal cycle that changes the temperature up and down in a cyclic manner.

204. The method according to claim 203, wherein the thermal cycle is for nucleic acid amplification using polymerase chain reaction.

205. The method according to claim 204, further comprising: changing the temperature of the sample for nucleic acid isothermal amplification.

206. The method according to claim 205, wherein the area of at least a portion of the sample and the radiation absorption layer is greater than the uniform thickness.

207. The device according to claim 190, wherein the radiation absorption layer comprises: a disk-coupled dot pillar antenna array, a silicon interlayer, graphene, a superlattice.

208. The device according to claim 190, wherein the radiation absorption layer comprises a carbon nanostructure or a black nanostructure.

209. The device according to claim 190, wherein the radiation absorption layer is configured to: absorb radiant energy.

210. The device according to claim 209, wherein the radiation absorption layer is configured to: radiate energy in the form of heat after absorbing radiant energy.

211. The device according to claim 190, wherein the radiation absorption layer is located below the sample layer and in direct contact with the sample layer.

212. The device according to claim 190, wherein the radiation absorption layer is configured to: absorb electromagnetic waves; the electromagnetic waves are selected from: radio waves, microwaves, infrared waves, visible light, ultraviolet waves, X-rays, gamma rays, and thermal radiation.

213. The device according to claim 190, wherein at least one plate does not block the radiation absorbed by the radiation absorption layer.

214. The device according to claim 186, wherein the thermal conductivity of one or both plates is lower than that of other parts.

215. The method according to claim 194, wherein the uniform thickness of the sample layer is adjusted by one or more spacers fixed to one or two plates.

216. The device according to claim 186, wherein the sample is a pre-mixed polymerase chain reaction medium.

217. The method according to claim 194, wherein the device is configured to change the temperature of the sample to facilitate a PCR assay according to a predetermined program.

218. The device according to claim 186, wherein the device is configured to perform diagnostic tests, health monitoring, environmental tests, and / or forensic identification.

219. The device according to claim 186, wherein the device is configured to perform DNA amplification, DNA quantification, selective DNA separation, genetic analysis, tissue typing, oncogene identification, infectious disease testing, genetic fingerprint analysis, and / or paternity testing.

220. The device according to claim 186, wherein the sample layer is sealed laterally to reduce sample evaporation.

221. The system according to claim 193, wherein the controller is configured to: control the presence, intensity, wavelength, frequency, and / or angle of electromagnetic waves.

222. The system according to claim 221, further comprising: a thermometer configured to measure the temperature at or near the sample contact area and send a signal to the controller based on the measured temperature.

223. The system according to claim 222, wherein the thermometer is selected from: fiber optic thermometer, infrared thermometer, liquid crystal thermometer, pyrometer, quartz thermometer, silicon bandgap temperature sensor, temperature strip, thermistor, and thermocouple.

224. The system according to claim 223, wherein the controller is configured to control the presence, intensity, wavelength, frequency, and / or angle of the electromagnetic wave from the radiation source.

225. The system according to claim 224, wherein the radiation source and the radiation absorption layer are configured such that the electromagnetic wave causes an average heating rate of at least 10 °C / s; and removing the electromagnetic wave causes an average cooling rate of at least 5 °C / s.

226. The device according to claim 192, wherein the radiation source and the radiation absorption layer are configured to produce an average heating rate of at least 10 °C / s and an average cooling rate of at least 5 °C / s.

227. The method according to claim 194, wherein the radiation source and the radiation absorption layer are configured to: produce an average heating rate of at least 10 °C / s to achieve the initialization step, denaturation step, and / or extension / elongation step in the PCR process, and an average cooling rate of at least 5 °C / s to achieve the annealing step and / or final cooling step in the PCR process.

228. The method according to claim 194, wherein the sample comprises: template DNA, primer DNA, cations, polymerase, and buffer.

229. The method according to claim 194, wherein the step of pressing the plate into a closed configuration comprises pressing the plate with non-precise pressure.

230. The method according to claim 229, wherein the step of pressing the plate into a closed configuration comprises directly pressing the plate by hand.

231. The method according to claim 194, wherein the thickness variation of the layer with very uniform thickness is less than 10%.

232. The method according to claim 194, wherein the temperature change of the sample is a thermal cycle, and the thermal cycle is used for nucleic acid amplification using polymerase chain reaction, and the polymerase chain reaction is selected from: hot start PCR, nested PCR, touchdown PCR, reverse transcription PCR, RACE PCR, and digital PCR.

233. The method according to claim 194, wherein the temperature change of the sample is used for nucleic acid isothermal amplification, and the nucleic acid isothermal amplification is selected from: loop-mediated isothermal amplification, strand displacement amplification, helicase-dependent amplification, nicking enzyme amplification, rolling circle amplification, and recombinase polymerase amplification.

234. The device according to claim 8, further comprising: reagents selected from: DNA template, primer, DNA polymerase, deoxynucleoside triphosphate, divalent cation, monovalent cation, and buffer solution.

235. The device according to claim 8, wherein the spacer has a flat top.

236. The device according to claim 8, wherein, any one of the plates has a thickness of 50 µm or less.

237. A device for rapidly changing the temperature of a thin fluid sample layer, comprising: A first plate, a second plate, a radiation absorption layer, and a spacer, wherein: i. The first plate and the second plate are movable relative to each other into different configurations, including an open configuration and a closed configuration; ii. Each plate includes a sample contact area on its respective surface for contacting a fluid sample, wherein the temperature of at least a portion of the fluid sample needs to be rapidly changed; iii. The first plate and the second plate have a configuration for rapidly changing the temperature of the sample; iv. The spacer has a uniform predetermined height equal to or less than 200 microns; v. At least one of the spacers is located within the sample contact area; wherein in the open configuration, the two plates are partially or fully separated, the spacing between the plates is not adjusted by the spacer, and the sample is deposited on one or both plates; and wherein, in the closed configuration configured after the sample is deposited in the open configuration, the two plates are parallel, at least a portion of the sample is squeezed by the two plates into a layer having a uniform thickness and stagnant relative to the plates, wherein the uniform thickness of the layer is defined by the sample contact areas of the two plates and is adjusted by the plates and the spacer, and wherein the plates are configured to change the temperature of the sample at a rate of at least 10 °C / sec, wherein a radiation absorption layer is further included, the radiation absorption layer is close to at least a portion of the sample having a uniform thickness, and the area of at least a portion of the sample and the radiation absorption layer is greater than the uniform thickness; The spacer is used to adjust the distance between the first plate and the second plate, and the ratio ISD of the fourth power of the spacer distance ISD to the product of the thickness h and Young's modulus E of the flexible plate 4 / (hE) is less than or equal to 5×10 6 µm³ / GPa.

238. A system for rapidly changing the temperature of a thin fluid sample layer, comprising: i. The device according to claim 237, ii. A radiation source, wherein the radiation source is configured to radiate electromagnetic waves significantly absorbed by the radiation absorption layer; and iii. A controller configured to control the radiation source and rapidly change the temperature of the sample.

239. A method for rapidly changing the temperature of a thin fluid sample layer, comprising: i. Providing the system according to claim 238; ii. Depositing a fluid sample on one or both plates; iii. Pressing the plates into a closed configuration; and iv. Changing and maintaining the temperature of the sample layer by changing the presence, intensity, wavelength, frequency, and / or angle of the electromagnetic waves from the radiation source.

240. The method according to claim 239, wherein the temperature change of the sample is a thermal cycle of changing the temperature up and down in a cyclic manner.

241. The method according to claim 239, wherein the temperature change of the sample is a thermal cycle, and the thermal cycle is for nucleic acid amplification using polymerase chain reaction.

242. The method according to claim 239, wherein the temperature change of the sample is for nucleic acid isothermal amplification.

243. The method according to claim 239, the area of the sample and the radiation absorption layer is greater than the uniform thickness.

244. The device according to claim 237, wherein the radiation absorption layer includes a disk-coupled dot column antenna array, a silicon interlayer, graphene, a superlattice.

245. The device according to claim 237, wherein the radiation absorption layer includes a carbon nanostructure or a black nanostructure.

246. The device according to claim 237, wherein the radiation absorption layer is configured to absorb radiation energy.

247. The device according to claim 237, wherein the radiation absorption layer is configured to radiate energy in the form of heat after absorbing radiation energy.

248. The device according to claim 237, wherein the radiation absorption layer is located below the sample layer and in direct contact with the sample layer.

249. The device according to claim 237, wherein the radiation absorption layer is configured to absorb electromagnetic waves selected from radio waves, microwaves, infrared waves, visible light, ultraviolet waves, X-rays, gamma rays, and thermal radiation.

250. The device according to claim 237, wherein at least one of the plates does not block the radiation absorbed by the radiation absorption layer.

251. The device according to claim 237, wherein the thermal conductivity of the first plate or / and the second plate is lower than that of other parts.

252. The method according to claim 239, wherein the uniform thickness of the sample layer is adjusted by one or more spacers fixed to one or two plates.

253. The method according to claim 239, wherein the sample is a premixed polymerase chain reaction medium.

254. The device according to claim 237, wherein the device is configured to change the temperature of the sample by promoting a PCR assay according to a predetermined program.

255. The device according to claim 237, wherein the device is configured to perform diagnostic tests, health monitoring, environmental tests, and / or forensic identification.

256. The device according to claim 237, wherein the device is configured to perform DNA amplification, DNA quantification, selective DNA separation, genetic analysis, tissue typing, oncogene identification, infectious disease testing, genetic fingerprint analysis, and / or paternity testing.

257. The method according to claim 239, wherein the sample layer is sealed laterally to reduce sample evaporation.

258. The system according to claim 238, wherein the controller is configured to control the presence, intensity, wavelength, frequency, and / or angle of the electromagnetic wave.

259. The system according to claim 238, further comprising: a thermometer configured to measure the temperature at or near the sample contact area and send a signal to the controller based on the measured temperature.

260. The system according to claim 259, wherein the thermometer is selected from: fiber optic thermometer, infrared thermometer, liquid crystal thermometer, pyrometer, quartz thermometer, silicon bandgap temperature sensor, temperature strip, thermistor, and thermocouple.

261. The system according to claim 238, wherein the radiation source and the radiation absorption layer are configured such that the electromagnetic wave causes an average heating rate of at least 10 °C / s; and removing the electromagnetic wave causes an average cooling rate of at least 5 °C / s.

262. The system according to claim 238, wherein the radiation source and the radiation absorption layer are configured to generate an average heating rate of at least 10 °C / s to achieve the initialization step, denaturation step, and / or extension / elongation step during the PCR process, and an average cooling rate of at least 5 °C / s to achieve the annealing step and / or final cooling step during the PCR process.

263. The method according to claim 19, wherein the first plate or the second plate further comprises a light absorption layer disposed on the heating / cooling layer, and wherein the light absorption layer has an average light absorption rate of at least 30%.

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