Cell analysis in body fluids, especially blood

Abstract

Apparatus and methods for analyzing analytes such as white blood cells in a liquid sample are disclosed.

Description

[0001] Cross - reference to related applications

[0002] This application claims the benefit of priority of U.S. Provisional Patent Application No. 62 / 764,887, filed on Aug. 16, 2018, and U.S. Provisional Patent Application No. 62 / 719,201, filed on Aug. 17, 2018. The entire contents of the above provisional patent applications are incorporated herein by reference in their entirety. The entire disclosures of any publications or patent documents mentioned herein are incorporated by reference in their entirety. Field of the invention

[0003] The present invention particularly relates to devices and methods for performing biological and chemical assays, such as but not limited to assays related to white blood cell analysis. Background art

[0004] In biological and chemical analysis (e.g., diagnostic tests), it is often necessary to quickly and simply measure and / or detect analytes in a sample or a portion of a sample. The present invention provides devices and methods for achieving these goals. Brief description of the drawings

[0005] 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 the drawings, some of the drawings are not drawn to scale. In the drawings presenting experimental data points, the lines connecting the data points are for guiding the observation of the data only and have no other purpose.

[0006] Figure 1 Illustrates an embodiment of QMAX (Q: Quantification); M: Magnification; A: Reagent addition; X: Acceleration; also known as a Compression - Regulated Open - Flow (CROF) device, which includes a first plate and a second plate. FIG. (A) shows a perspective view of the plates in an open configuration when the plates are separated; FIG. (B) shows a perspective view and a cross - sectional view of depositing a sample on the first plate in the open configuration; FIG. (C) is a perspective view and a cross - sectional view of the QMAX device in a closed configuration.

[0007] Figure 2 Illustrates a comparison of white blood cell (WBC) count accuracy, field of view (FoV), and QMAX gap (thickness of the sample layer). FIG. (A) shows a graph of WBC count accuracy versus QMAX gap size, with an effective FoV of 4 mm 2 , 16 mm 2 , 36 mm 2 , 64 mm 2 and 100 mm 2 ; FIG. (B) shows a graph of WBC count accuracy FoV with QMAX gap sizes of 2 μm, 3 μm, 5 μm, 6.2 μm, 10 μm, and 30 μm.

[0008] Figure 3 Figure (A) shows a graph comparing the percentage of WBC missing counts with the QMAX gap sizes (thickness of the sample layer) of 2μm, 5μm, 10μm, and 30μm. Figure (B) shows a graph of the QMAX transmittance at a wavelength of 500nm (which is close to the fluorescence of WBC) versus the QMAX gap size for the QMAX transmittance at a wavelength of 500nm (which is close to the fluorescence of WBC).

[0009] Figure 4 Shows the theoretical calculation of the self - overlap rate of WBC cells with respect to the QMAX gap.

[0010] Figure 5 Shows a schematic exploded view of an optical adapter device for attaching a QMAX device to a mobile communication device.

[0011] Figure 6 Shows a schematic cross - sectional view of the details of a system that can be used to test a sample in a fluorescence illumination mode, and specifically shows the optical adapter.

[0012] Figure 7 Shows (a) a photograph of the QMAX device and (b) a photograph of the QMAX device and the adapter on a smart phone.

[0013] Figure 8 Shows (a) a bright - field image of HgB in the device at a wavelength of approximately 520nm, (b) a bright - field image of WBC in the device at an excitation of approximately 490nm and an emission of over 500nm, (c) a bright - field image of RBC in the device, (d) a bright - field image of WBC and PLT in the device at an excitation of approximately 490nm and an emission of over 500nm, where the interior of whole blood is photographed by an optical system based on an iPhone.

[0014] Figure 9 Shows exemplary HgB, WBC, RBC, PLT analysis results of a whole - blood sample using the QMAX device and compares them with a commercial hematology counter such as the Horiba Pentra 60C. The results show that the device and method have better accuracy compared to commercial machines.

[0015] Detailed description of exemplary embodimentsThe following detailed description shows some embodiments of the present invention by way of example and not limitation. The section headings and any subtitles used here are for organizational purposes only and should not be construed as limiting the subject matter described in any way. The content under the section headings and / or sub - headings is not limited to the section headings and / or sub - headings, but applies to the entire description of the present invention.

[0016] Any reference to a publication is to its disclosure prior to the filing date and should not be construed as an admission that this claim is not entitled to antedate such publication by virtue of prior invention. In addition, the provided publication date may be different from the actual publication date, which may need to be independently confirmed.

[0017] Among other things, the present invention provides devices, systems, and methods for performing biological and chemical assays using QMAX cards.

[0018] Exemplary embodiments disclosed herein can be combined with biological / chemical devices and assays, including but not limited to those disclosed, described, and / or mentioned in Application PCT / US2016 / 046437, which is hereby incorporated herein by reference in its entirety.

[0019] The embodiments in these applications, which are hereby incorporated by reference, can be considered to be combined with each other or as a single invention, rather than as discrete and independent documents.

[0020] In addition, the exemplary embodiments disclosed herein are applicable to embodiments including but not limited to the following: biological / chemical assays, QMAX cards and systems, QMAX having hinges, notches, grooved edges, and sliders, assays and devices having uniform sample thickness, smartphone detection systems, cloud computing designs, various detection methods, labels, capture agents, and detection agents, analytes, diseases, applications, and samples; various embodiments are disclosed, described, and / or referenced in the above applications, all of which are hereby incorporated by reference in their entirety.

[0021] Example of a QMAX device with a hinge (QMAX card)

[0022] Figure 1An embodiment of a general QMAX (Q: Quantification; M: Amplification; A: Reagent addition; X: Acceleration; also known as Compression-Regulated Open Flow (CROF) device) is shown. The general QMAX device includes a first plate 10 and a second plate 2. In particular, Figure (A) shows a perspective view of the first plate 10 and the second plate 20, where the first plate has spacers. However, it should be noted that the spacers can also be fixed on the second plate 20 (not shown) or on both the first plate 10 and the second plate 20 (not shown). Figure (B) shows a perspective view and a cross-sectional view of depositing a sample 90 on the first plate 10 in an open configuration. However, it should be noted that the sample 90 can also be deposited on the second plate 20 (not shown), or on both the first plate 10 and the second plate 20 (not shown). Figure (C) shows (i) using the first plate 10 and the second plate 20 to distribute the sample 90 (the sample flows between the inner surfaces of the plates) and reduce the sample thickness, and (ii) using the spacers and the plates to adjust the sample thickness in the closed configuration of the QMAX device. The inner surface of each plate has one or more binding sites and / or storage sites (not shown).

[0023] In some embodiments, the spacer 40 has a predetermined uniform height and a predetermined uniform spacer pitch. In the closed configuration, as shown in Figure (C) of Figure 1 , the spacing between the plates and thus the thickness of the sample 90 are adjusted by the spacer 40. In some embodiments, the uniform thickness of the sample 90 is substantially similar to the uniform height of the spacer 40. It should be noted that although Figure 1 shows the spacer 40 fixed on one of the plates, in some embodiments, the spacer is not fixed. For example, in certain embodiments, the spacer is mixed with the sample so that when the sample is compressed into a thin layer, the spacer, as a rigid bead or particle with uniform size, adjusts the thickness of the sample layer.

[0024] QMAX assay

[0025] In biological and chemical assays (i.e., tests), devices and / or methods that simplify assay operations or accelerate assay speed are generally of great value.

[0026] In QMAX (Q: Quantification; M, Amplification; A, Reagent addition; X: Acceleration; also known as Compression-Regulated Open Flow (CROF) assay platform), the QMAX card uses two plates to manipulate the sample into a thin layer (e.g., by compression) as shown in Figure 1 . In certain embodiments, the plate manipulation requires multiple changes in the relative positions of the two plates (referred to as: plate configuration) by hand or other external forces. The QMAX card needs to be designed to make the manual operation easy and fast.

[0027] In the QMAX measurement, one of the plate configurations is an open configuration where the two plates are completely or partially separated (the spacing between the plates is not controlled by spacers) and a sample can be deposited. Another configuration is a closed configuration where at least a portion of the sample deposited in the open configuration is compressed between the two plates into a layer of very uniform thickness, the uniform thickness of which is defined by the inner surfaces of the plates and adjusted by the plates and spacers.

[0028] In the QMAX measurement operation, the operator needs to first place the two plates in the open configuration ready for sample deposition, then deposit the sample on one or both plates, and finally close the plates to the closed position. In some embodiments, the two plates of the QMAX card are initially on top of each other and need to be separated to enter the open configuration for sample deposition. When one of the plates is a thin plastic film (175 μm thick PMA), this separation is difficult to perform by hand. The present invention aims to provide an apparatus and method for making the operation of certain measurements (such as QMAX card measurements) easy and fast.

[0029] In some embodiments, the QMAX apparatus includes a hinge that connects two or more plates such that the plates can open and close in a manner similar to a book.

[0030] In certain embodiments, the hinge is configured such that the hinge can self - hold the angle between the plates after adjustment.

[0031] In certain embodiments, the hinge is configured such that the material of the hinge can hold the QMAX card in the closed configuration so that the entire QMAX card can be slid into and out of the card slot without causing accidental separation of the two plates.

[0032] Another aspect of the present invention is to provide an opening mechanism, such as but not limited to a notch on the edge of the plate or a strip attached to the plate, to make it easier for the user to manipulate the positioning of the plate, such as but not limited to separating the plates by hand.

[0033] Another aspect of the present invention is to provide a hinge that can control the rotation of more than two plates.

[0034] The term "Compression Open Flow (COF)" refers to a method of changing the shape of a flowable sample deposited on a plate by: (i) placing another plate on top of at least a portion of the sample, and (ii) then compressing the sample between the two plates by pushing the two plates towards each other; wherein the compression reduces the thickness of at least a portion of the sample and causes the sample to flow into the open space between the plates. The term "Compression Regulated Open Flow" or "CROF" (or "Self-Calibrating Compression Open Flow" or "SCOF" or "SCCOF") (also known as QMAX) refers to a specific type of COF, where the final thickness of part or all of the sample after compression is "regulated" by a spacer, where the spacer is placed between the two plates. Here, CROF devices can be used interchangeably with QMAX devices.

[0035] Unless otherwise specified, the term "spacer" or "stopper" refers to a mechanical object that sets a limit on the minimum spacing between two plates when placed between them, which limit can be reached when the two plates are compressed together. That is, during compression, the spacer will stop the relative movement of the two plates to prevent the plate spacing from becoming less than a preset (i.e., predetermined) value.

[0036] The terms "the spacer has a predetermined height" and "the spacer has a predetermined spacer spacing" respectively mean that the values of the spacer height and the spacer spacing are known prior to the QMAX process. If the values of the spacer height and the spacer spacing are not known prior to the QMAX process, then the values of the spacer height and the spacer spacing are not predetermined. For example, in the case where beads are sprayed as spacers onto a plate, where the beads land at random positions on the plate, the distance between the spacers is not predetermined. Another example of a non-predetermined spacer-to-spacer distance is when the spacer moves during the QMAX process.

[0037] In the QMAX process, the term "the spacer is fixed to its corresponding plate" means that the spacer is connected to a position on the plate and remains connected to that position during the QMAX process (i.e., the position of the spacer on the corresponding plate does not change). An example of "the spacer is fixed to its corresponding plate" is when the spacer is integrally made from a piece of the plate material and the position of the spacer relative to the plate surface does not change during the QMAX process. An example of "the spacer is not fixed to its corresponding plate" is when the spacer is adhered to the plate by an adhesive, but during the use of the plate, during the QMAX process, the adhesive cannot hold the spacer in its original position on the plate surface and the spacer moves away from its original position on the plate surface.

[0038] In the QMAX process, the term "open configuration" of two plates refers to a structure where the two plates are either partially or completely separated and the spacing between the plates is not regulated by a spacer

[0039] The "closed configuration" of the two plates in QMAX processing refers to the configuration in which the plates face each other, the relevant volume of the spacer and the sample is between the plates, and the relevant spacing between the plates and thus the thickness of the relevant volume of the sample are adjusted by the plates and the spacer, where the relevant volume is at least a part of the entire volume of the sample.

[0040] In QMAX processing, the term "the sample thickness is adjusted by the plates and the spacer" means that, for given conditions of the plates, the sample, the spacer, and the plate compression method, the thickness of at least one port of the sample in the closed configuration of the plates can be predetermined according to the properties of the spacer and the plates.

[0041] In a QMAX device, the term "inner surface" or "sample surface" of a plate refers to the surface of the plate that contacts the sample, and the other surface of the plate (not contacting the sample) is called the "outer surface".

[0042] Unless otherwise specified, the term "height" or "thickness" of an object in the QMAX process refers to the dimension of the object in the direction perpendicular to the plate surface. For example, the spacer height is the dimension of the spacer in the direction perpendicular to the plate surface, and the spacer height and the spacer thickness mean the same thing.

[0043] Unless otherwise specified, the term "area" of an object in the QMAX process refers to the area of the object parallel to the plate surface. For example, the spacer area is the spacer area parallel to the plate surface.

[0044] The term QMAX device refers to a device that performs the QMAX (e.g., CROF) process on a sample and has or does not have a hinge connecting two plates.

[0045] The terms "QMAX device with a hinge" and "QMAX card" are interchangeable.

[0046] The terms "angle self-maintain", "angle self-maintaining", or "rotation angle self-maintaining" refer to the property of a hinge that substantially maintains the angle between two plates after the external force that moves the plates from an initial angle to an angle is removed from the plates.

[0047] QMAX device and assay for cell counting

[0048] The QMAX device can be used to analyze fluid samples, such as but not limited to biological fluid samples. In some embodiments, the QMAX device is used to analyze blood samples. For example, in certain embodiments, the QMAX device is used to measure the amount of certain analytes, such as red blood cells (RBCs), white blood cells (WBCs), and / or count certain subtypes of blood cells. In certain embodiments, the QMAX device can be used for WBC counting. In certain embodiments, staining reagents can be used to label cells and structures, such as but not limited to RBCs, WBCs (including WBC subtypes), and platelets.

[0049] As Figure 1 shown, various parameters of the QMAX device can vary based on a particular test. For example, in some embodiments, the spacer height is less than 0.2μm, 0.5μm, 0.8μm, 1μm, 1.2μm, 1.5μm, 1.8μm, 2μm, 3μm, 4μm, 5μm, 6μm, 7μm, 8μm, 9μm, 10μm, 11μm, 12μm, 13μm, 14μm, 15μm, 16μm, 17μm, 18μm, 19μm, 20μm, 25μm, 30μm, 35μmμm, 40μm, 45μm, 50μm, 60μm, 70μm, 75μm, 80μm, 90μm, 100μm, 125μm, 150μm, 175μm, 200μm, 250μm, 300μm, 350μm, 400μm, 450μm, 500μm, 600μm, 700μm, 800μm, 900μm, 1mm, 2mm, 3mm, 4mm, 5mm, 10mm, or within a range between any two of the said values. In the closed configuration, the uniform thickness of the sample layer is substantially the same as the gap between the QMAX plates, and the gap between the QMAX plates is substantially the same as the spacer height. Therefore, the description of the spacer height also applies to the thickness of the sample layer and the QMAX gap, and vice versa.

[0050] In some embodiments of the QMAX assay, the sample is deposited onto one or two plates in an open configuration; then the plates are pressed into a closed configuration such that at least a portion of the sample is compressed into a layer of very uniform thickness that is stationary on the plates and defined by the inner surfaces of the plates. In some embodiments, the analyte in the sample is measured. In certain embodiments, the analyte is a cell type that can be counted. For example, in certain embodiments, the sample is a blood sample and the analyte is red blood cells; in certain embodiments, the sample is a blood sample and the analyte is white blood cells; in certain embodiments, the sample is a blood sample and the analyte is a white blood cell subtype (including neutrophils, eosinophils, basophils, lymphocytes, and monocytes).

[0051] In some embodiments, when the QMAX device is in a closed configuration, a camera can be used to capture an image of the sample layer. In certain embodiments, the camera can have a field of view (FoV), which is defined as the area within which the camera can capture an image of the sample. In certain embodiments, the camera is part of a device, such as but not limited to a mobile device. In certain embodiments, the mobile device is a smart phone, a tablet computer, or a laptop computer. In some embodiments, the mobile device is a mobile communication device, such as a smart phone. In certain embodiments, the camera has one lens; in certain embodiments, the camera has two lenses that are aligned parallel to each other.

[0052] In some embodiments, different spacer heights (and thus different sample thicknesses and QMAX gaps) can affect the accuracy of counting certain cells, such as but not limited to white blood cells and subtypes of white blood cells. For example, for counting white blood cells (WBCs), the spacer height and FoV affect the accuracy and consistency of the counting results. With an acceptable level of consistency, the direct counting results can be adjusted to reflect the true number of cells, providing a basis for diagnosis and health guidance. In certain embodiments, one factor to consider is the consistency of the "miss" rate, which is the deviation of the result of the tested method from the real number, and this deviation is typically established using a well-defined and well-accepted method. It should also be noted that the methods disclosed herein can be applied not only to WBC counting but also to other assays.

[0053] The devices and methods of the present invention can be used to (1) count white blood cells, (b) count subtypes of white blood cells (including neutrophils, eosinophils, basophils, lymphocytes, and monocytes), and (3) differentiate white blood cells, wherein the device further comprises a spacer that adjusts the spacing between sample contact areas when the plate is in a closed configuration.

[0054] In some embodiments, the average thickness of the uniformly thick layer is in the range of 5.0 μm to 8.5 μm.

[0055] In some embodiments, the average thickness of the uniformly thick layer is in the range of 7.5 μm to 10.5 μm.

[0056] In some embodiments, the average thickness of the uniformly thick layer is in the range of 9.5 μm to 12.5 μm.

[0057] In some embodiments, the average thickness of the uniformly thick layer is in the range of 9.5 μm to 12.5 μm.

[0058] In some embodiments, the average thickness of the uniformly thick layer is in the range of 11.5 μm to 13.5 μm.

[0059] In some embodiments, the average thickness of the uniform thickness layer is in the range of 12.5 μm to 14.5 μm.

[0060] In some embodiments, the average thickness of the uniform thickness layer is in the range of 13.5 μm to 16 μm.

[0061] In some embodiments, the spacer height is in the range of 5.0 μm to 8.5 μm.

[0062] In some embodiments, the spacer height is in the range of 7.5 μm to 10.5 μm.

[0063] In some embodiments, the spacer height is in the range of 9.5 μm to 12.5 μm.

[0064] In some embodiments, the spacer height is in the range of 9.5 μm to 12.5 μm.

[0065] In some embodiments, the spacer height is in the range of 11.5 μm to 13.5 μm.

[0066] In some embodiments, the spacer height is in the range of 12.5 μm to 14.5 μm.

[0067] In some embodiments, the spacer height is in the range of 13.5 μm to 16 μm.

[0068] In some embodiments, the field of view for counting and differentiating WBC is 0.1 mm 2 , 10 mm 2 , 50 mm 2 , 100 mm 2 , or in the range between any two of said values.

[0069] In some embodiments, when the gap size of the QMAX device is about 10 μm, the FoV is greater than 36 mm 2 , whereby the WBC counting and differentiation accuracy is less than 5%.

[0070] In some embodiments, when the gap size of the device is 10 μm, the FoV is greater than 16 mm 2 , whereby the WBC counting and differentiation accuracy is less than 10%.

[0071] In some embodiments, when the gap size of the device is 10 μm, the FoV is greater than 2 mm 2 , whereby the WBC counting and differentiation accuracy is less than 20%.

[0072] In some embodiments, the field of view is from 0.1 mm 2 to 10 mm 2Within the range, the preferred gap size of the device is in the range of 10 μm to 30 μm, 30 μm to 50 μm, whereby the counting and discrimination accuracy is less than 10%.

[0073] In some embodiments, the field of view is from 0.1 mm 2 to 10 mm 2 Within the range, the preferred gap size of the device is in the range of 10 μm to 30 μm, whereby the counting and discrimination accuracy is less than 20%.

[0074] In some embodiments, the field of view is from 10 mm 2 to 50 mm 2 Within the range, the preferred gap size of the device is in the range of 5 μm to 30 μm, 10 μm to 30 μm, whereby the counting and discrimination accuracy is less than 10%.

[0075] In some embodiments, the field of view is from 10 mm 2 to 50 mm 2 Within the range, the preferred gap size of the device is in the range of 2 μm to 5 μm, 5 μm to 10 μm, 10 μm to 30 μm, whereby the counting and discrimination accuracy is less than 20%.

[0076] In some embodiments, the field of view is from 50 mm 2 to 100 mm 2 Within the range, the preferred gap size of the device is in the range of 2 μm to 5 μm, 5 μm to 10 μm, 10 μm to 30 μm, 30 μm to 50 μm, whereby the counting and discrimination accuracy is less than 10%.

[0077] In some embodiments, the preferred height range of the spacer is 2 μm to 5 μm, whereby the WBC missing count is less than 15%.

[0078] In some embodiments, the preferred height range of the spacer is 2 μm to 5 μm, 5 μm to 10 μm, whereby the WBC missing count is less than 30%.

[0079] In some embodiments, the preferred height range of the spacer is 2 μm to 5 μm, 5 μm to 10 μm, 10 μm to 30 μm, whereby the WBC missing count is less than 60%.

[0080] In some embodiments, the distance from the sample to the lens is in the range of 2 mm to 5 mm.

[0081] In some embodiments, the distance from the sample to the lens is in the range of 4 mm to 7 mm.

[0082] In some embodiments, the distance from the sample to the lens is in the range of 6 mm to 9 mm.

[0083] In some embodiments, the distance from the sample to the lens ranges from 8 mm to 11 mm.

[0084] In some embodiments, the distance from the sample to the lens ranges from 10 mm to 13 mm.

[0085] In some embodiments, the distance from the sample to the lens ranges from 12 mm to 15 mm.

[0086] Example of a QMAX device for white blood cell counting

[0087] Figure 2 Illustrates the comparison of white blood cell (WBC) counting accuracy, field of view (FoV), and QMAX gap (thickness of the sample layer). Undiluted blood is deposited on one or both plates of the QMAX device in an open configuration; the plates are pressed into a closed configuration such that at least part of the sample is compressed into a layer of uniform thickness; an image of the compressed sample is captured using a camera in a smartphone; and WBC counting is performed by image analysis.

[0088] Figure 2 Figure (A) shows a graph comparing WBC counting accuracy with QMAX gap size, with an effective FoV of 4 mm 2 、16 mm 2 、36 mm 2 、64 mm 2 and 100 mm 2 ; Figure (B) shows a graph of WBC counting accuracy FoV with QMAX gap sizes of 2 μm, 3 μm, 5 μm, 6.2 μm, 10 μm, and 30 μm. The results are also summarized in Table 1.

[0089] Table 1 - Comparison of WBC counting accuracy, field of view, and QMAX gap

[0090]

[0091] In this set of experiments, the first plate of the QMAX device is a 1 - mm - thick PMMA with printed acridine orange dye, and the second plate is an X - plate with spacers made on 175 - μm - thick PMMA, having a column size of 30×40 μm and a spacer distance of 80 μm. 1 μL of fresh blood without any anticoagulant was used in the test and deposited on the first plate. The counting accuracy is defined as the standard deviation of the number of counts for all fields on the card with a specific FoV. This counting accuracy represents the situation when fields with FoV in the sample layer are randomly selected for measurement, indicating how accurate the average number of all fields is. Generally, WBC counting is more accurate with a larger field of view and a larger QMAX gap. In essence, the counting accuracy here reflects the consistency of the method with a specific gap size and field of view.

[0092] Table 2 shows the comparison of WBC missing counts and correction factors with QMAX gaps. In this article, the missing count rate is defined as the percentage difference between the back-calculated WBC concentration (from the count number, count area, fill factor, void size) and the true WBC concentration of the sample (measured by a calibrated commercial blood machine).

[0093] Correction factor = 1 / (1 - missing count rate).

[0094] Table 2 - Comparison of WBC Missing Counts, Correction Factors and QMAX Gaps

[0095] QMAX Gap Size (μm) WBC Deficiency Count WBC Correction Factor 2 0％ 1 3 0％ 1 5 10％ 1.1 10 25％ 1.3 30 50％ 2.0

[0096] As shown in Table 2, the missing count rate increases with the increase of the gap size (and thus the spacer height and sample thickness). In addition, additional experiments show that the differentiated WBC (granulocyte, lymphocyte, monocyte) counts have a similar missing count rate to the total WBC count. Additionally, the WBC missing count rate is not affected by the field of view.

[0097] Figure 3 Figure (A) shows a graph comparing the percentage of WBC missing counts with QMAX gap sizes of 2μm, 5μm, 10μm and 30μm (thickness of the sample layer; spacer height). Figure 3 Figure (B) shows a graph comparing the QMAX transmittance with the QMAX gap size at a wavelength of 500nm (which is close to the fluorescence of WBC).

[0098] As Figure 3 shown in, in Figures (A) and (B), as the gap size is larger (thicker blood film), more WBCs are missing. One reason is that the fluorescence of WBCs is dimmed and blocked by RBCs with a thicker blood film, as shown by the transmittance vs. gap size in (b). Therefore, the larger the QMAX gap, the more WBC missing counts. However, as Figure 2 shown in Figures (A) and (B) in, the counting accuracy reflecting the consistency of the counts at a specific gap size and field of view is higher at larger gap sizes and larger fields of view respectively. Therefore, in some embodiments, certain gap sizes (and thus spacer heights) and / or field of view sizes can be selected to obtain an acceptable level of consistency and / or prevent a high level of missing counts.

[0099] Using the correction factor based on the missing count rate, the counting results can be adjusted to provide a more accurate and consistent quantity for medical and health purposes. In some embodiments, the final quantity is equal to the counting result multiplied by the correction factor. In certain embodiments, the correction factor can be obtained / calculated from Table 2 and / or Figure 3 obtained / calculated.

[0100] Figure 4 The self - overlap rate of WBC cells is shown relative to the QMAX gap calculation. The results are also shown in Table 3. Generally, when the gap size is large, especially greater than 30 μm, more WBCs overlap.

[0101] Table 3 - Comparison of QMAX gap size, WBC distance, and overlap rate

[0102]

[0103]

[0104] Exemplary embodiments with gaps of 8 to 12 μm

[0105] Experiments (see, for example Figures 2 - 4 ) show that for the measurement of WBCs in an undiluted blood sample, at a given field of view provided by a camera (such as a camera in a mobile phone), a spacer height of 5 to 15 μm provides more accurate results than a spacer height of 2 to 3 μm. In some embodiments, the QMAX device for WBC measurement has a spacer height of 5 to 15 μm. In certain embodiments, the QMAX device has a spacer height of 10 μm, and the same sample thickness uniformity can be achieved. In some embodiments, such a column height is advantageous for imaging and counting white blood cells in undiluted blood.

[0106] Exemplary embodiments of the optical adapter

[0107] In some embodiments, a QMAX device with a sample (e.g., in the form of a QMAX card) can be inserted into an adapter, which can be attached to a device containing a camera and / or an illumination source. In certain embodiments, the device is a mobile communication device, such as but not limited to a smart phone.

[0108] Figure 5 A schematic exploded view of an optical adapter device for attaching a QMAX device to a mobile communication device and for measuring an analyte in a sample is shown. Here, the optical adapter device 18 is in a system 19 that includes a mobile communication device (smart phone) 1.

[0109] The adapter 18 includes a holder shell 2 assembled above the upper part of the smart phone 1; an optical box 3, which is attached to the shell 2 and includes a socket slot 4, an optical chamber 3C, multiple tracks 6b and 6t that allow a rod 8 to slide in, and a rubber door 16, which is inserted into multiple grooves 4s to cover the socket slot 4. An optical plug 7 is installed on the top of the optical chamber 3C, where the exit aperture 7L and the entry aperture 7C communicate with the light source 1L and the camera 1C in the smart phone 1 (see Figure 6)Alignment. The lens 11 is mounted in the entry aperture 7C of the optical cartridge 7 and is configured such that the sample in the insertion socket slot 4 in the sample carrier 5 is within the working distance of the camera 1C (see Figure 6 ). The lens 11 is used to magnify the image of the sample captured by the camera 1C (see below Figure 6 ). The long-pass optical filter 12 is mounted on top of the lens 11 in the entry aperture 7C. A pair of right-angle mirrors 13 and 14 are mounted on the bottom of the optical chamber 3C and are configured such that the mirror 13 and the mirror 14 are aligned with the light source 1L and the camera 1C, respectively (see Figure 6 ). The operation of the mirrors 13 and 14 as bright-field illumination optics in the device 18 is described below in Figure 6 .

[0110] The rod 8 includes two horizontal rods: the upper horizontal rod contains the band-pass optical filter 15 mounted in the slot 8a, and the lower horizontal rod contains the light absorber 9 mounted on the horizontal plane 8b and the reflector 10 mounted on the inclined plane 8c. The operation of the optical filter 15, the light absorber 9, and the reflector 10 as fluorescence illumination optics in the device 18 is described in Figure 6 . The upper horizontal rod of the rod 8 slides along the track 6t in the box 3, and the lower horizontal rods 8b and 8c slide along the track 6b in the box 3. The rod 8 stops at two different positions in the box 3 to switch between the bright-field illumination optics and the fluorescence illumination optics. The rod 8 is fully inserted into the box 3 to switch the device 18 to work with the fluorescence illumination optics. The ball plug 17 is mounted on the side wall of the track 6t to stop the rod 8 at a predetermined position when the rod 8 is pulled out of the box 3 to switch the device 18 to work with the bright-field illumination optics.

[0111] Figure 6 A schematic cross-sectional view showing details of a system that can be used to test a sample in a fluorescence illumination mode is shown, and details of the optical adapter are specifically shown. The figure shows the functions of the elements described above with reference to Figure 5 . The rod 8 (shown in Figure 5The fully inserted device 18 positions the light absorber 9 and the tilted mirror 10 within the field of view of the camera 1C and the light source 1L, blocking the optical path between the light source 1L and the pair of mirrors 13 and 14. The bandpass optical filter 15 is located directly below the light source 1L. The light source 1L emits a light beam BF1 away from the smartphone 1. The optical filter 15 allows the light beam BF1 with a specific wavelength range matching the excitation wavelength of the fluorescent sample in the sample slide 5 to pass through. A portion of the light beam BF1 irradiates on the edge of the transparent sample slide 5, couples into the waveguide light beam BF3 traveling in the sample slide 5, and irradiates on the sample area under the lens 11. A portion of the light beam BF1 irradiates on the mirror 10. The tilted mirror 10 deflects the beam BF1 into the beam BF2 and irradiates the sample area in the sample slide 5 in reverse at a larger tilt angle directly below the lens 11. The remaining portion of the light beam BF1 with a large divergence angle (i.e., the light beam BF4) irradiates on the absorber 9 and is absorbed, such that the reflected light of the light beam BF4 does not enter the camera 1C at a small incident angle. The light from the sample area under the lens 11 passes through the lens 11 and is filtered by the long-pass filter 12, allowing only the light within the specified wavelength range emitted by the fluorescent sample in the sample slide 5 to enter the camera 1C to form an image. The smartphone 1 captures and processes the image to obtain some properties of the sample. The rubber door 16 is inserted into the device 18 to cover the sample slide 5, preventing ambient light from entering the device 18 and affecting the test.

[0112] In some embodiments, Figure 5 and 6 the adapter described in can be used to measure blood samples, such as undiluted whole blood samples. In certain embodiments, the analyte can be WBC, for which the rod 8 needs to be inserted for optimal reading. In some embodiments, the adapter comprises:

[0113] (a) An attachment member configured to attach the adapter to a device comprising a light source and a camera;

[0114] (b) A card slot configured to accommodate a sample card containing a liquid sample compressed into a uniformly thick layer, wherein when the sample card is inserted into the card slot, the sample is within the field of view of the camera and the light source;

[0115] (c) An optical filter configured to filter the light from the light source to form a first light beam with a specific wavelength range, wherein a portion of the first light beam irradiates on the edge of the sample card and travels in the sample card to irradiate the sample;

[0116] (d) A mirror configured to deflect a portion of the first light beam to form a second light beam that irradiates the sample in reverse at an inclined angle;

[0117] (e) An absorber configured to absorb the remaining portion of the first light beam with a divergence angle.

[0118] In some embodiments, a method for measuring an analyte, such as but not limited to WBC, in a liquid sample may comprise:

[0119] (a) Obtaining a liquid sample;

[0120] (b) Compressing at least a portion of the sample into a layer of uniform thickness using a sample card,

[0121] (c) Inserting the sample card into an adapter device configured to attach to a device comprising a light source and a camera;

[0122] (d) Irradiating the sample with light from the light source, wherein

[0123] i. The light is filtered by an optical filter of the adapter device to form a first light beam having a specific wavelength range, and a portion of the first light beam irradiates an edge of the sample card and travels through the sample card to irradiate the sample;

[0124] ii. A portion of the first light beam is deflected by a mirror of the adapter device to form a second light beam that irradiates the sample in a reverse direction at an inclined angle; and

[0125] iii. The remaining portion of the first light beam having a divergence angle is absorbed by an absorber of the adapter device.

[0126] In some embodiments, the method further comprises:

[0127] (a) Capturing an image of the sample in the layer of uniform thickness using a camera;

[0128] (b) Analyzing the image to enumerate the analyte in the image; and

[0129] (c) Calculating the concentration of the analyte in the sample based on the uniform thickness, the field of view of the camera, the number of analytes, and a predetermined correction factor;

[0130] wherein the field of view is the range of the field in which the camera captures an image;

[0131] wherein the correction factor is determined by an error counting ratio that depends on the field of view, the uniform thickness, and the nature of the analyte.

[0132] Exemplary embodiments for WBC measurement

[0133] For an apparatus or method embodiment of the present invention, the apparatus may further comprise a multi-reagent layer on one or two plates, the multi-reagent layer including an anti-adhesion reagent, a cell lysis reagent, a cell staining reagent, a release time control material, and any combination thereof.

[0134] In some embodiments, each reagent layer coated on the plate has a thickness of 10 nm, 100 nm, 200 nm, 500 nm, 1 μm, or within a range between any two of these values.

[0135] In some embodiments, the anti-adhesion agent comprises ethylenediaminetetraacetic acid (EDTA), disodium ethylenediaminetetraacetate, K2EDTA, or K3EDTA, or any combination thereof.

[0136] In some embodiments, the cell stain comprises Wright's stain (eosin, methylene blue), Giemsa stain (eosin, methylene blue, and azur B), May-Grünwald stain, Leishman's solution (“polychrome” methylene blue (i.e., demethylated to azurin) and eosin), erythrosin B stain (erythrosin B), and other fluorescent dyes, including but not limited to acridine orange dye, 3,3 - dioctadecyloxacarbocyanine (DiOC6), propidium iodide (PI), fluorescein isothiocyanate (FITC), and basic orange 21 (BO21) dye, ethidium bromide, lucigenin, and diamidinostilbene disulfonic acid derivatives, erythrosin B or trypan blue, Hoechst 33342, trihydrochloride, Trihydrate, DAPI (4,6 - diamidino - 2 - phenylindole, dihydrochloride), or any combination thereof.

[0137] In some embodiments, the cell lysing agent comprises ammonium chloride, sodium bicarbonate, ethylenediaminetetraacetic acid (EDTA), acetic acid, citric acid, or other acids and bases, or any combination thereof.

[0138] In some embodiments, the release time control material comprises albumin, carbomer, carboxymethyl cellulose, carrageenan, chitosan, dextrin, polyethylene glycol, polyvinylpyrrolidone, or polyvinyl alcohol, or any combination thereof.

[0139] In some embodiments of the method embodiments of the present invention, the RBC, platelets, or both in the sample are lysed before detecting and / or measuring WBC.

[0140] In some embodiments of the method embodiments of the present invention, the WBC, platelets, or both are lysed in the sample before detecting RBC.

[0141] In some embodiments of the method embodiments of the present invention, the RBC, WBC, or both are lysed in the sample before detecting PLT.

[0142] A set of other examples of the present invention

[0143] Other embodiments of the inventive subject matter according to the present disclosure are described in the paragraphs listed below.

[0144] Correction Factor and Field of View

[0145] A1. A method for analyzing an analyte in a liquid sample, comprising:

[0146] (a) obtaining a liquid sample;

[0147] (b) compressing at least a portion of the sample into a layer of uniform thickness,

[0148] (c) capturing an image of the sample in the layer of uniform thickness using a camera, wherein the image shows the analyte; and

[0149] (d) analyzing the image to enumerate the analyte in the image,

[0150] (e) calculating the concentration of the analyte in the sample based on the uniform thickness, the field of view of the camera, the analyte enumeration, and a predetermined correction factor;

[0151] wherein the field of view is the range of the field in which the camera captures the image;

[0152] wherein the correction factor is determined by an error count ratio that depends on the field of view, the uniform thickness, and the nature of the analyte.

[0153] Irradiation of WBC

[0154] B1. An adapter device for analyzing an analyte in a liquid sample, comprising:

[0155] (a) an attachment member configured to attach the adapter to a device comprising a light source and a camera;

[0156] (b) a card slot configured to receive a sample card containing a liquid sample compressed into a layer of uniform thickness, wherein when the sample card is inserted into the card slot, the sample is under the field of view of the camera and the light source;

[0157] (c) an optical filter configured to filter light from the light source to form a first light beam having a specific wavelength range, wherein a portion of the first light beam irradiates an edge of the sample card and travels in the sample card to irradiate the sample;

[0158] (d) a mirror configured to deflect a portion of the first light beam to form a second light beam that irradiates the sample in the reverse direction at an inclined angle;

[0159] (e) an absorber configured to absorb the remaining portion of the first light beam having a divergence angle.

[0160] B2. A method for analyzing an analyte in a liquid sample, comprising:

[0161] (a) obtaining a liquid sample;

[0162] (b) using a sample card to compress at least a portion of the sample into a layer of uniform thickness,

[0163] (c) Insert the sample card into the adapter device, which is configured to attach to a device comprising a light source and a camera;

[0164] (d) Irradiate the sample with light from the light source, wherein

[0165] i. The light is filtered by an optical filter of the adapter device to form a first light beam having a specific wavelength range, and a portion of the first light beam irradiates the edge of the sample card and travels through the sample card to irradiate the sample;

[0166] ii. A portion of the first light beam is deflected by a mirror of the adapter device to form a second light beam that irradiates the sample in a reverse direction at an inclined angle; and

[0167] iii. The remaining portion of the first light beam having a divergence angle is absorbed by an absorber of the adapter device.

[0168] B3. The kit according to embodiment 35A, further comprising:

[0169] (a) Capture an image of the sample in a layer of uniform thickness using the camera;

[0170] (b) Analyze the image to enumerate the analytes in the image; and

[0171] (c) Calculate the concentration of the analytes in the sample based on the uniform thickness, the field of view of the camera, the number of analytes, and a predetermined correction factor;

[0172] wherein the field of view is the range of the field in which the camera captures the image;

[0173] wherein the correction factor is determined by an error counting ratio, and the error counting ratio depends on the field of view, the uniform thickness, and the nature of the analytes.

[0174] Additional Features:

[0175] C1. The device or method according to any one of the above embodiments, wherein the liquid sample is a blood sample.

[0176] C2. The device or method according to any one of the above embodiments, wherein the analyte is a white blood cell (WBC).

[0177] C3. The device or method according to any one of the above embodiments, wherein the analyte is a WBC subtype.

[0178] C4. The device or method according to any one of the above embodiments, wherein the analyte is a neutrophil, an eosinophil, a basophil, a lymphocyte, or a monocyte.

[0179] C5. The device or method according to any one of the above embodiments, wherein a fluorescent label is used to analyze the analyte.

[0180] C6. The device or method according to any one of the above embodiments, wherein the uniform thickness is in the range of 5 to 30 μm.

[0181] C7. The device or method according to any one of the above embodiments, wherein the uniform thickness is in the range of 8 to 12 μm.

[0182] C8. The device or method according to any one of the above embodiments, wherein the uniform thickness is about 10 μm.

[0183] C9. The device or method according to any one of the above embodiments, wherein the field of view (FOV) is equal to or greater than 4 mm 2 。

[0184] C10. The device or method according to any one of the above embodiments, wherein the field of view (FOV) is equal to or greater than 16 mm 2 。

[0185] C11. The device or method according to any one of the above embodiments, wherein the field of view (FOV) is equal to or greater than 36 mm 2 。

[0186] C12. The device or method according to any one of the above embodiments, wherein the field of view (FOV) is equal to or greater than 64 mm 2 。

[0187] C13. The device or method according to any one of the above embodiments, wherein the field of view (FOV) is equal to or greater than 100 mm 2 。

[0188] C14. The device or method according to any one of the above embodiments, wherein when the sample thickness is 2 mm, the correction factor is 1, when the sample thickness is 3 mm, the correction factor is 1, when the sample thickness is 5 mm, the correction factor is 1.1, when the sample thickness is 10 mm, the correction factor is 1.3, and when the sample thickness is 30 mm, the correction factor is 2.0.

[0189] C15. The device or method according to any one of the above embodiments, wherein the analyte is labeled with a fluorescent label, and the wavelength range of the first light beam matches the excitation wavelength of the fluorescence of the labeled analyte.

[0190] C15. The device or method according to any one of the above embodiments, wherein the adapter device further includes a housing member.

[0191] C16. The device or method according to any one of the above embodiments, wherein the adapter device further comprises a rod that can be inserted into or withdrawn from the housing member.

[0192] C17. The device or method according to any one of the above embodiments, wherein the mirror and the absorber are mounted on the rod.

[0193] C18. The device or method according to any one of the above embodiments, wherein the adapter device comprises a card slot having a safety opening that allows a sample card to be inserted and prevents ambient light from entering the card slot.

[0194] WBC Analysis Device

[0195] AA1. A device for analyzing white blood cells in a blood sample, comprising:

[0196] A first plate, a second plate, and a spacer, wherein:

[0197] i. The plates can be moved relative to each other into different configurations;

[0198] ii. One or both of the plates are flexible;

[0199] iii. Each of the plates comprises an inner surface having a sample contact area for contacting the blood sample;

[0200] iv. One or both of the plates comprise a spacer permanently fixed to the sample contact area of the respective plate;

[0201] v. The spacer has:

[0202] (a) A predetermined substantially uniform height having a value selected in the range of 2 μm to 30 μm,

[0203] (b) The shape of a column having a substantially uniform cross-section and a flat top surface;

[0204] (c) The ratio of the width to the height is equal to or greater than 1;

[0205] (d) A predetermined fixed, non-random spacer pitch in the range of 10 μm to 200 μm (micrometers);

[0206] (e) A fill factor equal to 1% or greater, where the fill factor is the ratio of the spacer contact area (on the plate) to the total plate area; and

[0207] (f) The product of the fill factor and the Young's modulus is 2 MPa or greater.

[0208] One of the configurations is an open configuration, in which the two plates are partially or completely separated, the spacing between the plates is not adjusted by spacers, and the sample is deposited on one or both of the plates; and

[0209] The other of the configurations is a closed configuration, which is configured after the sample is deposited in the open configuration; and in the closed configuration, at least a portion of the sample is compressed into a layer with a very uniform thickness and is substantially stagnant relative to the plates, wherein the uniform thickness of the layer is limited by the sample contact area of the two plates and is adjusted by the plates and spacers.

[0210] AA2. An apparatus for analyzing white blood cells in a blood sample, comprising:

[0211] A first plate, a second plate, spacers, and an adapter, wherein:

[0212] i. The plates are movable relative to each other into different configurations;

[0213] ii. One or both of the plates are flexible;

[0214] Each of the plates includes an inner surface having a sample contact area for contacting a fluid sample.

[0215] iv. One or both of the plates include spacers permanently fixed to the sample contact area of the respective plate;

[0216] v. The spacers have:

[0217] (a) A predetermined substantially uniform height, which has a value selected in the range of 2 μm to 30 μm,

[0218] (b) The shape of a column, which has a substantially uniform cross-section and a flat top surface;

[0219] (c) The ratio of the width to the height is equal to or greater than 1;

[0220] (d) A predetermined fixed, non-random spacer spacing in the range of 10 μm to 200 μm;

[0221] (e) A packing factor equal to 1% or greater, where the packing factor is the ratio of the spacer contact area (on the plate) to the total plate area; and

[0222] (f) The product of the packing factor and the Young's modulus is 2 MPa or greater.

[0223] vi. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone having a camera, (c) a slot in the housing that allows (1) a plate in a closed configuration to slide into the slot, and (2) when the plate is in the slot, at least a portion of the sample area is less than 2 cm from the outer surface of the camera, and (d) an optical system in the housing configured to image at least a portion of the sample contact area by the camera;

[0224] 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 a spacer, and the sample is deposited on one or both of the plates; and

[0225] The other configuration is a closed configuration that is configured after the sample is deposited in the open configuration; and in the closed configuration, at least a portion of the sample is compressed into a layer of very uniform thickness and is substantially stationary relative to the plates, wherein the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the plates and the spacer.

[0226] AA3. A device for analyzing white blood cells in a blood sample, comprising:

[0227] A first plate, a second plate, a spacer, and an adapter, wherein:

[0228] i. The plates are movable relative to each other into different configurations;

[0229] ii. One or both of the plates are flexible;

[0230] iii. Each of the plates includes an inner surface having a sample contact area for contacting a fluid sample.

[0231] iv. One or both of the plates include spacers permanently fixed on the sample contact areas of the corresponding plates;

[0232] v. The spacer has:

[0233] (a) a predetermined substantially uniform height having a value selected in the range of 10 μm to 50 μm,

[0234] (b) the shape of a column having a substantially uniform cross-section and a flat top surface;

[0235] (c) the ratio of the width to the height is equal to or greater than 1;

[0236] (d) a predetermined fixed, non-random spacer spacing in the range of 10 μm to 200 μm;

[0237] (e) a fill factor equal to or greater than 3%, where the fill factor is the ratio of the spacer contact area (on the plate) to the total plate area; and

[0238] (f) the product of the fill factor and the Young's modulus is 2 MPa or greater.

[0239] vi. One or both of the plates contain a reagent coated on the sample contact area of the corresponding plate;

[0240] vii. The reagent has at least one of the following: (a) a component that stains WBC; (b) a component that makes RBCs distribute uniformly; (c) a component that lyses RBCs;

[0241] viii. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone with a camera, (c) a slot in the housing that allows (1) the plate in the closed configuration to slide into the slot, and (2) when the plate is in the slot, at least a part of the sample area is less than 2 cm from the outer surface of the camera, and (d) an optical system in the housing that is configured to image at least a part of the sample contact area by the camera;

[0242] where one of the configurations is an open configuration, in which the two plates are partially or completely separated, the spacing between the plates is not adjusted by the spacer, and the sample is deposited on one or both of the plates; and

[0243] where the other of the configurations is a closed configuration, which is configured after the sample is deposited in the open configuration; and in the closed configuration, at least a part of the sample is compressed into a layer with a very uniform thickness and is substantially stationary relative to the plates, where the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the plates and the spacer.

[0244] AA4. A method for analyzing white blood cells in a blood sample, comprising:

[0245] (a) obtaining a blood sample;

[0246] (b) obtaining the device AA1 or AA2 or AA3;

[0247] (c) when the plate is configured in the open configuration, depositing the blood sample on one or both of the plates,

[0248] (d) after (c), forcing the two plates to form a closed configuration; and

[0249] (e) when the plate is in the closed configuration, capturing an image of the sample in the layer with a uniform thickness; and

[0250] (f) Analyze the image to determine the concentration of white blood cells in the sample.

[0251] AA5. A method for counting white blood cells and subtypes (including neutrophils, eosinophils, basophils, lymphocytes, and monocytes) using a single device, comprising:

[0252] (a) Obtain a blood sample;

[0253] (b) Obtain the device according to any one of the above embodiments, wherein the spacer height is 5 μm to 40 μm,

[0254] (c) When the plate is configured in an open configuration, deposit the blood sample on one or both of the plates,

[0255] (d) After (c), force the two plates to form a closed configuration;

[0256] (e) When the plate is in the closed configuration, capture an image of the sample in a layer of uniform thickness; and

[0257] (f) Analyze the image to determine the respective numbers of white blood cells, neutrophils, lymphocytes, monocytes, eosinophils, and basophils by counting the number of cells in the image and analyzing the fluorescence color (meaning emission wavelength range) and shape of each white blood cell.

[0258] BB1. The device or method according to any one of the above embodiments, wherein the blood sample is undiluted.

[0259] BB2. The device or method according to any one of the above embodiments, wherein the staining and shape of white blood cells provide differentiation of fluorescence color, structure, and size of white blood cells and their subtypes, as well as white blood cell differentiation.

[0260] The device or method according to any one of the above embodiments, wherein the fluorescence color (meaning emission wavelength range) is used for WBC counting and differentiation;

[0261] The device or method according to any one of the above embodiments, wherein the size is used for WBC counting and differentiation;

[0262] The device or method according to any one of the above embodiments, wherein the structure of WBC is used for WBC counting and differentiation;

[0263] The device or method according to any one of the above embodiments, wherein both the color (meaning emission wavelength range) and structure of WBC are used for WBC counting and differentiation;

[0264] The device or method according to any one of the above embodiments, wherein the color (meaning emission wavelength range) of white blood cells is distinguished through the red, green, and blue channels of the image.

[0265] The device or method according to any of the above embodiments, wherein the color of white blood cells (meaning the emission wavelength range) is distinguished by a filter at different wavelengths before the camera.

[0266] The device or method according to any of the above embodiments, wherein the color of white blood cells (meaning the emission wavelength range) is distinguished by a filter at different wavelengths before the light source.

[0267] The device or method according to any of the above embodiments, wherein the color of white blood cells (meaning the emission wavelength range) comes from WBC staining with a chemical such as acridine orange dye.

[0268] The device or method according to any of the above embodiments, wherein the color of white blood cells (meaning the emission wavelength range) comes from WBC staining with different chemicals.

[0269] The device or method according to any of the above embodiments, wherein white blood cells are stained with propidium iodide (PI), fluorescein isothiocyanate (FITC), and basic orange (BOM); PI is a red nucleic acid dye that stains DNA and shows red fluorescence; FITC stains cytoplasmic proteins and shows green fluorescence; BO21 is a green nucleic acid dye that stains DNA and shows green fluorescence; the combination of these three fluorescent dyes can distinguish different types of white blood cells, such as lymphocytes, monocytes, neutrophils, and eosinophils.

[0270] The device or method according to any of the above embodiments, wherein the shape of white blood cells is analyzed by machine learning.

[0271] The device or method according to any of the above embodiments, wherein the color (meaning the emission wavelength range) and shape of white blood cells are analyzed by machine learning.

[0272] BB3. The device or method according to any of the above embodiments, wherein the device further comprises a multi-reagent layer on one or two plates, and the multi-reagent layer includes an anti-adhesion, cell lysis, cell staining, release time control material layer, or a combination thereof.

[0273] CC1. The device or method according to any of the above embodiments, wherein the column height is in the range of 5 μm to 15 μm.

[0274] CC2. The device or method according to any of the above embodiments, wherein the column height is in the range of 8 μm to 12 μm.

[0275] CC3. The device or method according to any of the above embodiments, wherein the column height is about 10 μm.

[0276] CC3. The device or method according to any one of the above embodiments, wherein the column height is about 30 μm.

[0277] CC4. The device or method according to any one of the above embodiments, wherein the device is configured to count white blood cells.

[0278] CC5. The device or method according to any one of the above embodiments, wherein the device is configured to count white blood cell subtypes (including neutrophils, eosinophils, basophils, lymphocytes, and monocytes).

[0279] CC6. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 7.5 μm to 10.5 μm.

[0280] CC7. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 9.5 μm to 12.5 μm.

[0281] CC8. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 11.5 μm to 13.5 μm.

[0282] CC9. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 12.5 μm to 14.5 μm.

[0283] CC10. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 13.5 μm to 15 μm.

[0284] CC10. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 15 μm to 18 μm.

[0285] CC10. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 18 μm to 25 μm.

[0286] CC10. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 25 μm to 30 μm.

[0287] CC10. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 30 μm to 35 μm.

[0288] CC10. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 35 μm to 40 μm.

[0289] CC10. The device or method according to any one of the above embodiments, wherein the spacer height is in the range of 40 μm to 50 μm.

[0290] CC11. The device or method according to any one of the above embodiments, wherein the field of view for counting and differentiating WBC is 0.1 mm 2 , 10 mm 2 , 50 mm 2 , 100 mm 2 , or in the range between any two of said values.

[0291] CC12. The device or method according to any one of the above embodiments, wherein when the gap size of the device is 10 μm, the FoV is greater than 36 mm 2 , whereby the WBC counting and differentiation accuracy is less than 5%.

[0292] CC13. The device or method according to any one of the above embodiments, wherein when the gap size of the device is 10 μm, the FoV is greater than 16 mm 2 , whereby the WBC counting and differentiation accuracy is less than 10%.

[0293] CC14. The device or method according to any one of the above embodiments, wherein when the gap size of the device is 10 μm, the FoV is greater than 2 mm 2 , whereby the WBC counting and differentiation accuracy is less than 20%.

[0294] CC15. The device or method according to any one of the above embodiments, wherein the field of view is 0.1 mm 2 to 10 mm 2 , and the preferred gap size of the device is in the range of 10 μm to 30 μm, 30 μm to 50 μm, whereby the counting and differentiation accuracy is less than 10%.

[0295] CC16. The device or method according to any one of the above embodiments, wherein the field of view is 0.1 mm 2 to 10 mm 2 , and the preferred gap size of the device is in the range of 10 μm to 30 μm, whereby the counting and differentiation accuracy is less than 20%.

[0296] CC17. The device or method according to any one of the above embodiments, wherein, wherein the field of view is 10 mm 2 to 50 mm 2 , and the preferred gap size of the device is in the range of 5 μm to 30 μm, 10 μm to 30 μm, whereby the counting and differentiation accuracy is less than 10%.

[0297] CC18. The device or method according to any one of the above embodiments, wherein the field of view is 10 mm 2 to 50 mm 2 , and the preferred gap size of the device is in the range of 2 μm to 5 μm, 5 μm to 10 μm, 10 μm to 30 μm, whereby the counting and discrimination accuracy is less than 20%.

[0298] CC19. The device or method according to any one of the above embodiments, wherein the field of view is 50 mm 2 to 100 mm 2 of the field of view, and the preferred gap size of the device is in the range of 2 μm to 5 μm, 5 μm to 10 μm, 10 μm to 30 μm, 30 μm to 50 μm, whereby the counting and differentiation accuracy is less than 10%.

[0299] CC20. The device or method according to any one of the above embodiments, wherein the height of the spacer is in the range of 2 μm to 5 μm, whereby the WBC missing count is less than 15%.

[0300] CC21. The device or method according to any one of the above embodiments, wherein the height of the spacer is in the range of 2 μm to 5 μm, 5 μm to 10 μm, whereby the WBC missing count is less than 30%.

[0301] CC22. The device or method according to any one of the above embodiments, wherein the preferred range of the height of the spacer is 2 μm to 5 μm, 5 μm to 10 μm, 10 μm to 30 μm, whereby the WBC missing count is less than 60%.

[0302] CC23. The device or method according to any one of the above embodiments, wherein the distance from the sample to the objective lens is in the range of 2 mm to 5 mm.

[0303] CC24. The device or method according to any one of the above embodiments, wherein the distance from the sample to the objective lens is in the range of 4 mm to 7 mm.

[0304] CC25. The device or method according to any one of the above embodiments, wherein the distance from the sample to the objective lens is in the range of 6 mm to 9 mm.

[0305] CC26. The device or method according to any one of the above embodiments, wherein the distance from the sample to the objective lens is in the range of 8 mm to 11 mm.

[0306] CC27. The device or method according to any one of the above embodiments, wherein the distance from the sample to the objective lens is in the range of 10 mm to 13 mm.

[0307] CC28. The apparatus or method according to any one of the above embodiments, wherein the distance from the sample to the objective lens is in the range of 12 mm to 15 mm.

[0308] Other examples of blood cell counting

[0309] An apparatus for analyzing white blood cells in a blood sample, comprising:

[0310] A first plate, a second plate, a spacer, and an adapter, wherein:

[0311] i. The plates are movable relative to each other into different configurations;

[0312] ii. One or both of the plates are flexible;

[0313] iii. Each of the plates includes an inner surface having a sample contact area for contacting a fluid sample.

[0314] iv. One or both of the plates include spacers permanently fixed to the sample contact areas of the respective plates;

[0315] v. The spacers have:

[0316] (a) A predetermined substantially uniform height having a value selected in the range of 10 μm to 50 μm,

[0317] (b) The shape of a column having a substantially uniform cross-section and a flat top surface;

[0318] (c) The ratio of the width to the height is equal to or greater than 1;

[0319] (d) A predetermined fixed, non-random spacer spacing in the range of 10 μm to 200 μm;

[0320] (e) A fill factor equal to 3% or greater, where the fill factor is the ratio of the spacer contact area (on the plate) to the total plate area; and

[0321] (f) The product of the fill factor and the Young's modulus is 2 MPa or greater.

[0322] vi. One or both of the plates include a reagent coated on the sample contact area of the respective plate;

[0323] vii. The reagent has at least one of the following: (a) a component that stains WBCs; (b) a component that equalizes the distribution of RBCs; (c) a component that lyses RBCs; (d) a component that dilutes the blood;

[0324] viii. The adapter includes: (a) a housing, (b) an attachment member on the housing that permits the adapter to be attached to a mobile phone having a camera, (c) a slot in the housing that permits (1) a plate in a closed configuration to slide into the slot, and (2) when the plate is in the slot, at least a portion of the sample area is less than 2 cm from the outer surface of the camera, and (d) an optical system in the housing configured to image at least a portion of the sample contact area by the camera;

[0325] 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 a spacer, and the sample is deposited on one or both of the plates; and

[0326] The other of the configurations is a closed configuration, which is configured after the sample is deposited in the open configuration; and in the closed configuration, at least a portion of the sample is compressed into a layer of very uniform thickness and is substantially stationary relative to the plates, wherein the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the plates and the spacer.

[0327] An apparatus for analyzing hemoglobin in a blood sample, comprising:

[0328] A first plate, a second plate, a spacer, and an adapter, wherein:

[0329] i. The plates are movable relative to each other into different configurations;

[0330] ii. One or both of the plates are flexible;

[0331] iii. Each of the plates includes an inner surface having a sample contact area for contacting a fluid sample.

[0332] iv. One or both of the plates include spacers permanently fixed to the sample contact areas of the respective plates;

[0333] v. The spacer has:

[0334] (a) a predetermined substantially uniform height having a value selected in the range of 10 μm to 50 μm,

[0335] (b) the shape of a column having a substantially uniform cross-section and a flat top surface;

[0336] (c) the ratio of the width to the height is equal to or greater than 1;

[0337] (d) a predetermined fixed, non-random spacer spacing in the range of 10 μm to 200 μm;

[0338] (e) A fill factor equal to or greater than 3%, where the fill factor is the ratio of the spacer contact area (on the plate) to the total plate area; and

[0339] (f) The product of the fill factor and the Young's modulus is 2 MPa or greater.

[0340] vi. One or both of the plates contain a reagent coated on the sample contact area of the corresponding plate;

[0341] vii. The reagent has at least one of the following: (a) a component that makes the RBCs distribute evenly; (b) a component that lyses the RBCs; (c) a component that dilutes the blood;

[0342] viii. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone with a camera, (c) a slot in the housing that allows (1) the plate in the closed configuration to slide into the slot, and (2) when the plate is in the slot, at least a part of the sample area is less than 2 cm from the outer surface of the camera;

[0343] where 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 spacer, and the sample is deposited on one or both of the plates; and

[0344] where the other of the configurations is a closed configuration, which is configured after the sample is deposited in the open configuration; and in the closed configuration, at least a part of the sample is compressed into a layer with a very uniform thickness by the two plates and is substantially stagnant relative to the plates, where the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the plates and the spacer.

[0345] An apparatus for analyzing red blood cells in a blood sample, comprising:

[0346] A first plate, a second plate, a spacer, and an adapter, where:

[0347] i. The plates are movable relative to each other into different configurations;

[0348] ii. One or both of the plates are flexible;

[0349] iii. Each of the plates includes an inner surface having a sample contact area for contacting a fluid sample.

[0350] iv. One or both of the plates include spacers permanently fixed on the sample contact areas of the corresponding plates;

[0351] v. The spacer has:

[0352] (a) A predetermined substantially uniform height having a value selected in the range of 1.5 μm to 8 μm,

[0353] (b) The shape of the columns, which have a substantially uniform cross-section and a flat top surface;

[0354] (c) The ratio of the width to the height is equal to or greater than 1;

[0355] (d) A predetermined fixed, non-random spacer pitch in the range of 10 μm to 200 μm;

[0356] (e) A fill factor equal to 3% or greater, where the fill factor is the ratio of the spacer contact area (on the plate) to the total plate area; and

[0357] (f) The product of the fill factor and the Young's modulus is 2 MPa or greater.

[0358] vi. One or both of the plates contain a reagent coated on the sample contact area of the respective plate;

[0359] vii. The reagent has at least one of the following: (a) a component that equalizes the distribution of RBCs; (b) a component that reduces the aggregation of RBCs; (c) a component that stains RBCs; (d) a component that dilutes the blood;

[0360] viii. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone having a camera, (c) a slot in the housing that allows (1) the plate in the closed configuration to slide into the slot, and (2) when the plate is in the slot, at least a portion of the sample area is less than 2 cm from the outer surface of the camera, and (d) an optical system in the housing configured to image at least a portion of the sample contact area by the camera;

[0361] 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 the spacers, and the sample is deposited on one or both of the plates; and

[0362] wherein the other of the configurations is a closed configuration that is configured after the sample is deposited in the open configuration; and in the closed configuration, at least a portion of the sample is compressed by the two plates into a layer of very uniform thickness and is substantially stationary relative to the plates, wherein the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the plates and the spacers.

[0363] An apparatus for analyzing platelets in a blood sample, comprising:

[0364] A first plate, a second plate, spacers, and an adapter, wherein:

[0365] i. The plates can move relative to each other into different configurations;

[0366] ii. One or both of the plates are flexible;

[0367] iii. Each of the plates includes an inner surface having a sample contact region for contacting a fluid sample.

[0368] iv. One or both of the plates include spacers permanently fixed on the sample contact regions of the respective plates;

[0369] v. The spacers have:

[0370] (a) A predetermined substantially uniform height having a value selected in the range of 1.5 μm to 30 μm,

[0371] (b) The shape of a column having a substantially uniform cross-section and a flat top surface;

[0372] (c) A ratio of width to said height equal to or greater than 1;

[0373] (d) A predetermined fixed, non-random spacer pitch in the range of 10 μm to 200 μm;

[0374] (e) A fill factor equal to 3% or greater, where the fill factor is the ratio of the spacer contact area (on the plate) to the total plate area; and

[0375] (f) The product of the fill factor and the Young's modulus is 2 MPa or greater.

[0376] vi. One or both of the plates include a reagent coated on the sample contact regions of the respective plates;

[0377] vii. The reagent has at least one of the following: (a) a component for equalizing the PLT distribution; (b) a component for reducing PLT aggregation; (c) a component for staining PLT; (d) a component for diluting blood;

[0378] viii. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone having a camera, (c) a slot in the housing that allows (1) the plates in the closed configuration to slide into the slot, and (2) when the plates are in the slot, at least a portion of the sample region is less than 2 cm from the outer surface of the camera;

[0379] 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 the spacers, and the sample is deposited on one or both of the plates; and

[0380] Another one of the configurations is a closed configuration, which is configured after the sample is deposited in the open configuration; and in the closed configuration, at least a portion of the sample is compressed between two plates into a layer with a very uniform thickness and is substantially stagnant relative to the plates, wherein the uniform thickness of the layer is limited by the sample contact area of the two plates and is adjusted by the plates and the spacers.

[0381] Spacer

[0382] According to the present invention, the spacing between the two plates and thus the sample thickness are controlled by using spacers.

[0383] Spacer height. In some embodiments, all spacers have the same predetermined height. In some embodiments, the spacers have different predetermined heights. In some embodiments, the spacers can be divided into groups or regions, where each group or region has its own spacer height. In certain embodiments, the predetermined height of the spacer is the average height of the spacer. In some embodiments, the spacers have approximately the same height. In some embodiments, a certain percentage number of spacers have the same height.

[0384] The height of the spacer is selected based on the desired adjusted spacing between the plates and / or the adjusted final sample thickness and the residual 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, less than 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 the range between any two of the said values.

[0385] The spacer height, the spacing between the plates and / or the sample thickness is 1 nm to 100 nm in a 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.

[0386] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 1.5 μm and 2.5 μm.

[0387] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 2.5 μm and 4 μm.

[0388] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 4 μm and 6 μm.

[0389] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 6 μm and 10 μm.

[0390] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 10 μm and 15 μm.

[0391] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 15 μm and 25 μm.

[0392] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 25 μm and 35 μm.

[0393] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 35 μm and 50 μm.

[0394] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 50 μm and 100 μm.

[0395] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 100 μm and 150 μm.

[0396] In a preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness are between 150 μm and 200 μm.

[0397] The spacer height is related to and limited by the incident light source power density when testing a whole blood sample.

[0398] In a preferred embodiment, at an incident light source power of 0.1 W / cm 2 to 5 W / cm 2 the spacer height, the spacing between the plates and / or the sample thickness are less than 2 μm, less than 5 μm, and less than 10 μm in a preferred embodiment.

[0399] In a preferred embodiment, at an incident light source power of 0.1 W / cm 2 to 5 W / cm2 In one preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness is less than 10 μm, less than 20 μm, less than 30 μm.

[0400] In one preferred embodiment, when the incident light source power is 0.1 W / cm 2 to 5 W / cm 2 In one preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness is less than 30 μm, less than 40 μm, less than 50 μm.

[0401] In one preferred embodiment, when the incident light source power is 5 W / cm 2 to 50 W / cm 2 In one preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness is less than 10 μm, less than 20 μm, less than 30 μm.

[0402] In one preferred embodiment, when the incident light source power is 5 W / cm 2 to 50 W / cm 2 In one preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness is less than 30 μm, less than 40 μm, less than 50 μm.

[0403] In one preferred embodiment, when the incident light source power is 5 W / cm 2 to 50 W / cm 2 In one preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness is less than 50 μm, less than 100 μm, less than 150 μm, less than 200 μm.

[0404] In one preferred embodiment, when the incident light source power is 50 W / cm 2 to 500 W / cm 2 In one preferred embodiment, the spacer height, the spacing between the plates, and / or the sample thickness is less than 50 μm, less than 100 μm, less than 150 μm, less than 200 μm.

[0405] In some embodiments, the spacer height is precisely controlled. The relative precision 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 two of said values.

[0406] In some embodiments, the spacer height, the spacing between the plates, and / or the sample thickness is: (i) equal to or slightly greater than the minimum size of the analyte, or (ii) equal to or slightly greater than the maximum size of the analyte. "Slightly greater than" means approximately 1% to 5% larger, and any value between the two values.

[0407] 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.

[0408] 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 spacing of the plates in the relevant area is 2 μm (equal to the minimum size) in one embodiment, 2.2 μm in another embodiment, or 3 μm (50% larger than the minimum size) in another embodiment, or 5 μm in another embodiment, but less than the maximum size of the red blood cells. Such embodiments have certain advantages in blood cell counting. In one embodiment, for red blood cell counting, by making the inner surface spacing 2 μm or 6 μm and any value between the two values, an undiluted whole blood sample is confined within this spacing; on average, each red blood cell (RBC) does not overlap with others, thus allowing for visually accurate counting of red blood cells. (Too much overlap between RBCs may lead to serious errors in counting).

[0409] For example, white blood cells have a size of 5 μm to 20 μm. In an embodiment of the present invention, the spacer is selected such that the inner surface spacing of the plates in the relevant area is 5 μm (equal to the minimum size) in one embodiment, 10 μm in another embodiment, or 30 μm (50% larger than the minimum size) in another embodiment, or 5 μm in another embodiment, but less than the maximum size of the red blood cells. Such embodiments have certain advantages in blood cell counting. In one embodiment, for white blood cell counting, by making the inner surface spacing 5 μm or 30 μm and any value between the two values, an undiluted whole blood sample is confined within this spacing, thus allowing for visually accurate counting of red blood cells.

[0410] In some embodiments, the spacer height, the spacing between the plates, and / or the sample thickness is: (i) equal to or slightly less than the minimum size of the analyte, or (ii) equal to or slightly less than the maximum size of the analyte. "Slightly less than" means it is approximately 1% to 5% smaller, and any value between the two values.

[0411] 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.

[0412] In the present invention, in some embodiments, the plates and spacers are used not only to adjust the thickness of the sample, but also to adjust the orientation and / or surface density of the analyte / entity in the sample when the plate is in the closed configuration. When the plate is in the closed configuration, the thinner thickness of the sample results in fewer analytes / entities per surface area (i.e., lower surface concentration).

[0413] Spacer lateral dimensions. For open spacers, the lateral dimensions can be characterized by its lateral dimensions (sometimes referred to as width) in two orthogonal directions, x and y. The lateral dimensions of the spacer are the same or different in each direction. In some embodiments, the lateral dimension in each direction (x or y) is 1 nm or less, 3 nm or less, 5 nm or less, 7 nm or less, 10 nm or less, 20 nm or less, 30 nm or less, 40 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, or 500 μm or less, or within a range between any two of the aforementioned values.

[0414] In some embodiments, the lateral dimension of the spacer is from 5 μm to 10 μm.

[0415] In some embodiments, the lateral dimension of the spacer is from 10 μm to 15 μm.

[0416] In some embodiments, the lateral dimension of the spacer is from 15 μm to 20 μm.

[0417] In some embodiments, the lateral dimension of the spacer is from 20 μm to 25 μm.

[0418] In some embodiments, the lateral dimension of the spacer is from 25 μm to 30 μm.

[0419] In some embodiments, the lateral dimension of the spacer is from 30 μm to 40 μm.

[0420] In some embodiments, the lateral dimension of the spacer is from 40 μm to 50 μm.

[0421] In some embodiments, the lateral dimension of the spacer is from 50 μm to 70 μm.

[0422] In some embodiments, the lateral dimension of the spacer is from 70 μm to 90 μm.

[0423] In some embodiments, the lateral dimension of the spacer is from 90 μm to 120 μm.

[0424] In some embodiments, the lateral dimension of the spacer is between 20 times and 40 times the center wavelength of the incident light.

[0425] In some embodiments, the lateral dimension of the spacer is between 40 times and 80 times the center wavelength of the incident light.

[0426] In some embodiments, the lateral dimension of the spacer is between 80 times and 120 times the center wavelength of the incident light.

[0427] In some embodiments, the lateral dimension of the spacer is between 120 times and 80 times the center wavelength of the incident light.

[0428] 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 in the range between any two of these values. In some embodiments, different ratios are used to adjust the sample flow direction; the larger the ratio, the more the flow is along one direction (the larger dimension direction).

[0429] In some embodiments, the different lateral dimensions of the spacer in the x and y directions are used for (a) using the spacer as a scale marker to indicate the orientation of the plate, (b) using the spacer to create more sample flow in a preferred direction, or both.

[0430] In a preferred embodiment, the period, width, and height of the spacer are substantially the same. In some embodiments, all spacers have the same shape and size. In some embodiments, the spacers have different lateral dimensions.

[0431] For a closed spacer, in some embodiments, the internal lateral shape and size 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 size are selected based on the required strength to support the pressure of the liquid against the spacer and the compression pressure of the pressing plate.

[0432] In certain embodiments, the aspect ratio of the height of the column 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 in the range between any two of these values.

[0433] Spacer spacing. The spacers can be single spacers 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 a fixed-spacing, non-fixed-spacing array or a fixed-spacing array at some positions on the plate, while being non-fixed-spacing at other positions.

[0434] In some embodiments, the spaced-apart 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.

[0435] In some embodiments, the spacer spacing of the spacer array is fixed-spacing (i.e., uniform spacer spacing) in at least one direction of the array. In some embodiments, the spacer spacing is configured to improve the uniformity between the plate spacings in the closed configuration.

[0436] In some embodiments, the distance between adjacent spacers (i.e., spacer spacing) 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 of the said values.

[0437] In certain embodiments, the spacer spacing 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 any range between the said values. In certain embodiments, the spacer spacing 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 any range between the said values.

[0438] The distance between adjacent spacers (i.e., spacer spacing) 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 any range between the said values; or in certain embodiments, at most 80%, 100%, 200%, 400%, or within any range between any two of the said values.

[0439] Obviously, in order to maintain a given sample thickness variation between two adjacent spacers, when using a more flexible plate, a closer spacer spacing is required.

[0440] In a preferred embodiment, the spacer pitch is between 20 μm and 50 μm.

[0441] In a preferred embodiment, the spacer pitch is between 50 μm and 80 μm.

[0442] In a preferred embodiment, the spacer pitch is between 80 μm and 100 μm.

[0443] In a preferred embodiment, the spacer pitch is between 100 μm and 150 μm.

[0444] In a preferred embodiment, the spacer pitch is between 150 μm and 200 μm.

[0445] In a preferred embodiment, the spacer pitch is between 200 μm and 250 μm.

[0446] In a preferred embodiment, the spacer pitch is between 250 μm and 300 μm.

[0447] In a preferred embodiment, the spacer pitch is between 300 μm and 400 μm.

[0448] In a preferred embodiment, the spacer pitch is between 400 μm and 500 μm.

[0449] In a preferred embodiment, the spacers are a regularly spaced square array, where the spacers are pillars with a height of 2 μm to 6 μm, an average lateral dimension of 10 μm to 40 μm, and a spacer pitch of 1 μm to 100 μm.

[0450] In a preferred embodiment, the spacers are a regularly spaced square array, where the spacers are pillars with a height of 2 μm to 6 μm, an average lateral dimension of 10 μm to 50 μm, and a spacer pitch of 100 μm to 250 μm.

[0451] In a preferred embodiment, the spacers are a regularly spaced square array, where the spacers are pillars with a height of 10 μm to 50 μm, an average lateral dimension of 20 μm to 50 μm, and a spacer pitch of 1 μm to 100 μm.

[0452] In a preferred embodiment, the spacers are a regularly spaced square array, where the spacers are pillars with a height of 10 μm to 50 μm, an average lateral dimension of 20 μm to 50 μm, and a spacer pitch of 100 μm to 250 μm.

[0453] The spacing 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, 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.

[0454] Spacer density. The spacers are arranged on the respective plates at the following surface densities: per μm 2 greater than 1, per 10 μm 2 greater than 1, per 100 μm 2 greater than 1, per 500 μm 2 greater than 1, per 1000 μm 2 greater than 1, per 5000 μm 2 greater than 1, per 0.01 mm 2 greater than 1, per 0.1 mm 2 greater than 1, per 1 mm 2 greater than 1, per 5 mm 2 greater than 1, per 10 mm 2 greater than 1, per 100 mm 2 greater than 1, per 1000 mm 2 greater than 1, per 10000 mm 2 greater than 1, or within the range between any two of said values. In some embodiments, the spacers have a density of at least 1 / mm 2 , at least 10 / mm 2 , at least 50 / mm 2 , at least 100 / mm 2 , at least 1,000 / mm 2 or at least 10,000 / mm 2 of density.

[0455] The spacer area filling factor is defined as the ratio of the spacer area to the total area or the ratio of the spacer spacing to the width. In some embodiments, the filling factor is at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, or within the range between any two of said values. In certain embodiments, the filling factor is at least 2.3%.

[0456] In a device comprising two plates and spacers, the fourth power of the spacer spacing (ISD) divided by the thickness (h) and Young's modulus (E) of the flexible plate (ISD 4 / (hE)) is 5×10 6 μm 3 / GPa or less.

[0457] In a device comprising two plates and spacers, the fourth power of the spacer spacing (ISD) divided by the thickness (h) and Young's modulus (E) of the flexible plate (ISD 4 / (hE)) is 5×10 5 μm 3 / GPa or less.

[0458] In a device comprising two plates and spacers, the spacers have a columnar shape, a substantially flat top surface, a predetermined substantially uniform height, and a predetermined constant spacer spacing, the spacer spacing being at least about 2 times larger than the size of the analyte, wherein the Young's modulus of the spacer multiplied by the fill factor of the spacer is equal to or greater than 2 MPa, wherein the fill factor is the ratio of the spacer contact area to the total plate area, and wherein for each spacer, the ratio of the lateral dimension of the spacer to its height is at least 1 (one).

[0459] In a device comprising two plates and spacers, the spacers have a columnar shape, a substantially flat top surface, a predetermined substantially uniform height, and a predetermined constant spacer spacing, the spacer spacing being at least about 2 times larger than the size of the analyte, wherein the Young's modulus of the spacer multiplied by the fill factor of the spacer is equal to or greater than 2 MPa, wherein the fill factor is the ratio of the spacer contact area to the total plate area, and wherein for each spacer, the ratio of the lateral dimension of the spacer to its height is at least 1 (one), wherein the fourth power of the spacer spacing (ISD) divided by the thickness (h) and Young's modulus (E) of the flexible plate (ISD 4 / (hE)) is 5x10 6 μm 3 / GPa or less.

[0460] In a device comprising two plates and spacers, the ratio of the spacer spacing of the spacers to the average width of the spacers is 2 or greater, and the fill factor of the spacers multiplied by the Young's modulus of the spacers is 2 MPa or greater.

[0461] The area, width and length of the card

[0462] For the device, kit, system or method according to any one of the above embodiments, wherein the area of any one of the plates depends on the specific application.

[0463] The device, kit, system or method according to any one of the above embodiments, wherein the area of at least one of the plates 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, 10000 cm 2 or less, or within the range between any two of said values.

[0464] The device, kit, system or method according to any one of the above embodiments, wherein the area of at least one plate of the QMAX card is in the range of 500 to 1000 mm 2 ;

[0465] The device, kit, system or method according to any one of the above embodiments, wherein the area of one plate is about 600 mm 2 , and the area of another plate is about 750 mm 2 .

[0466] The device, kit, system or method according to any one of the above embodiments, wherein at least one plate of the QMAX card is 1 mm or less, 5 mm or less, 10 mm or less, 15 mm or less, 20 mm or less, 25 mm or less, 30 mm or less, 35 mm or less, 40 mm or less, 45 mm or less, 50 mm or less, 100 mm or less, 200 mm or less, 500 mm or less, 1000 mm or less, 5000 mm or less, or within the range between any two of said values.

[0467] The device, kit, system or method according to any one of the above embodiments, wherein the width of at least one plate of the QMAX card is in the range of 20 mm to 30 mm.

[0468] The device, kit, system or method according to any one of the above embodiments, wherein the width of one plate is about 22 mm, and the width of another plate is about 24 mm.

[0469] The device, kit, system or method according to any one of the above embodiments, wherein at least one plate of the QMAX card is 1 mm or less, 5 mm or less, 10 mm or less, 15 mm or less, 20 mm or less, 25 mm or less, 30 mm or less, 35 mm or less, 40 mm or less, 45 mm or less, 50 mm or less, 100 mm or less, 200 mm or less, 500 mm or less, 1000 mm or less, 5000 mm or less, or within a range between any two of these values.

[0470] The device, kit, system or method according to any one of the above embodiments, wherein the length of at least one plate of the QMAX card is in the range of 20 to 40 mm;

[0471] The device, kit, system or method according to any one of the above embodiments, wherein the length of one plate is about 27 mm and the length of the other plate is about 32 mm.

[0472] The device, kit, system or method according to any one of the above embodiments, wherein the length of one plate is about 27 mm and the width of this plate is about 22 mm.

[0473] The device, kit, system or method according to any one of the above embodiments, wherein the length of one plate is about 32 mm and the width of this plate is about 24 mm.

[0474] The device, kit, system or method according to any one of the above embodiments, wherein the length of one plate is about 27 mm and the length of the other plate is about 32 mm, and the width of one plate is about 22 mm and the width of the other plate is about 24 mm.

[0475] Shape of the card

[0476] The device, kit, system or method according to any one of the above embodiments, wherein the shapes of the two plates are circular, oval, rectangular, triangular, polygonal, annular, or any superimposition of these shapes.

[0477] The device, kit, system or method according to any one of the above embodiments, wherein the two (or more) plates of the QMAX card can have the same dimensions and / or shapes, or different dimensions and / or shapes.

[0478] The device, kit, system or method according to any one of the above embodiments, wherein at least one of the two (or more) plates of the QMAX card has rounded corners for user safety considerations, and the diameter of the rounded corners is 100 μm or less, 200 μm or less, 500 μm or less, 1 mm or less, 2 mm or less, 5 mm or less, 10 mm or less, 50 mm or less, or within a range between any two of these values.

[0479] The device, kit, system or method according to any one of the above embodiments, wherein the plate can have any shape, preferably a shape that allows for a compressive open flow of the sample and adjustment of the sample thickness.

[0480] The device, kit, system or method according to any one of the above embodiments, wherein a special shape of the plate is advantageous.

[0481] The thickness of the card

[0482] The device, kit, system or method according to any one of the above embodiments, wherein the thickness, width and / or length of the two (or more) plates of the QMAX card can be the same or different.

[0483] The device, kit, system or method according to any one of the above embodiments, wherein the average thickness of at least one of the plates 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 (micrometers) 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 (millimeters) 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

[0484] The device, kit, system or method according to any one of the above embodiments, wherein the thickness of at least one of the plates is in the range of 0.5 mm to 1.5 mm.

[0485] The device, kit, system or method according to any one of the above embodiments, wherein the thickness of at least one of the plates is about 1 mm.

[0486] The device, kit, system or method according to any one of the above embodiments, wherein the thickness of at least one of the plates is 0.15 mm to 0.2 mm.

[0487] The device, kit, system or method according to any one of the above embodiments, wherein the thickness of at least one of the plates is about 0.175 mm.

[0488] The device, kit, system or method according to any one of the above embodiments, wherein the thickness of at least one of the plates is in the range of 0.01 mm to 0.15 mm.

[0489] The device, kit, system or method according to any one of the above embodiments, wherein the thickness of at least one of the plates is about 0.025 mm.

[0490] The device, kit, system or method according to any one of the above embodiments, wherein the thickness of at least one of the plates is about 0.05 mm.

[0491] The device, kit, system or method according to any one of the above embodiments, wherein the thickness of at least one of the plates is about 0.1 mm.

[0492] The device, kit, system or method according to any one of the above embodiments, wherein the thickness of any one of the plates is non-uniform on the plate.

[0493] The device, kit, system or method according to any one of the above embodiments, wherein different plate thicknesses at different positions can be used to control plate bending, folding, sample thickness adjustment, etc.

[0494] Notch

[0495] The device, kit, system or method according to any one of the above embodiments, wherein one or more notches are on one or more sides of one of the plates for easily peeling off another plate and separating the two plates.

[0496] The device, kit, system or method according to any one of the above embodiments, wherein the shape of the notch is circular, oval, rectangular, triangular, polygonal, annular or any superposition of these shapes.

[0497] The device, kit, system or method according to any one of the above embodiments, wherein the size of the notch is 1 mm 2 (square millimeters) or less, 10 mm 2 or less, 25 mm 2 or less, 50 mm 2 or less, 75 mm 2 or less or within the range between any two of the said values.

[0498] The device, kit, system or method according to any one of the above embodiments, wherein the area of each notch on the QMAX card is in the range of 10 to 30 mm 2 within the range.

[0499] The device, kit, system or method according to any one of the above embodiments, wherein the notch is a semi-circular shape with a diameter of 3 to 6 mm.

[0500] The device, kit, system or method according to any one of the above embodiments, wherein the notch has a width of 3 mm and a length of 6 mm.

[0501] The device, kit, system or method according to any one of the above embodiments, wherein the notch is located on the short-width side of the thicker plate.

[0502] The device, kit, system or method according to any one of the above embodiments, wherein two notches are located on the two long-width sides of the thicker plate.

[0503] hinge

[0504] The device, kit, system or method according to any one of the above embodiments, wherein the size of the hinge is variable and can be adjusted according to the size of the plate and the specific requirements of the device application.

[0505] The device, kit, system or method according to any one of the above embodiments, wherein the shape of the hinge is circular, oval, rectangular, triangular, polygonal, annular, or any superposition of these shapes.

[0506] The device, kit, system or method according to any one of the above embodiments, wherein the length of the hinge joint is less than 1 mm, 2 mm, 3 mm, 4 mm, 5 mm, 10 mm, 15 mm, 20 mm, 25 mm, 30 mm, 40 mm, 50 mm, 100 mm, 200 mm or 500 mm, or within the range between any two of said values.

[0507] The device, kit, system or method according to any one of the above embodiments, wherein the length of the hinge joint is about 20 mm.

[0508] The device, kit, system or method according to any one of the above embodiments, wherein the width of the hinge joint is less than 1 mm, 2 mm, 3 mm, 4 mm, 5 mm, 10 mm, 15 mm, 20 mm, 25 mm, 30 mm, 40 mm, 50 mm, 100 mm, 200 mm or 500 mm, or within the range between any two of said values.

[0509] The device, kit, system or method according to any one of the above embodiments, wherein the width of the hinge joint is about 6 mm.

[0510] The device, kit, system, or method according to any one of the above embodiments, wherein the length of the hinge joint is about 20 mm, and the width of the hinge joint is about 6 mm.

[0511] The device, kit, system, or method according to any one of the above embodiments, wherein the ratio of the length of the hinge joint to the length of the edge of the plate that the hinge joint aligns with is less than 1.5, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, 0.1, 0.05, or within the range between any two of these values.

[0512] The device, kit, system, or method according to any one of the above embodiments, wherein the ratio of the length of the hinge joint to the length of the edge of the plate that the hinge joint 36 aligns with is 1, indicating that the hinge joint completely covers the hinge edge.

[0513] The device, kit, system, or method according to any one of the above embodiments, wherein the total area of the hinge is less than 1 mm 2 , 5 mm 2 , 10 mm 2 , 20 mm 2 , 30 mm 2 , 40 mm 2 , 50 mm 2 , 100 mm 2 , 200 mm 2 , 500 mm 2 , or within the range between any two of these values.

[0514] The device, kit, system, or method according to any one of the above embodiments, wherein the width of the hinge joint is about 120 mm 2 .

[0515] The device, kit, system, or method according to any one of the above embodiments, wherein the ratio of the total size of the hinge to the total size of one of the plates is less than 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, 0.1, 0.05, 0.01, or within the range between any two of these values.

[0516] The device, kit, system, or method according to any one of the above embodiments, wherein the ratio of the total size of the hinge to the total size of one of the plates is about 0.16 to 0.20.

[0517] The device, kit, system, or method according to any one of the above embodiments, wherein the different layers of the hinge have the same or different thicknesses.

[0518] The device, kit, system or method according to any one of the above embodiments, wherein the thickness of any layer of the hinge is 0.1 μm, 1 μm, 2 μm, 3 μm, 5 μm, 10 μm, 20 μm, 30 μm, 50 μm, 100 μm, 200 μm, 300 μm, 500 μm, 1 mm, 2 mm, and within the range between any two of these values.

[0519] The device, kit, system or method according to any one of the above embodiments, wherein any layer in the layers of the hinge has a thickness within the range of 25 μm to 50 μm.

[0520] The device, kit, system or method according to any one of the above embodiments, wherein any layer in the layers of the hinge has a thickness within the range of 50 μm to 75 μm.

[0521] The device, kit, system or method according to any one of the above embodiments, wherein the hinge has a thickness of about 68 μm.

[0522] The device, kit, system or method according to any one of the above embodiments, wherein the length of the hinge joint is about 20 mm, the width of the hinge joint is about 6 mm, and the thickness of the hinge joint is about 68 μm.

[0523] Socket slot

[0524] The device, kit, system or method according to any one of the above embodiments, wherein the area of the receiving region of the socket slot or the lateral region covered by the sliding track is greater than or equal to the area of the QMAX device.

[0525] The device, kit, system or method according to any one of the above embodiments, wherein the shape of the receiving region of the socket slot is circular, oval, rectangular, triangular, polygonal, annular, or any superposition of these shapes.

[0526] The device, kit, system or method according to any one of the above embodiments, wherein the average gap size of the sliding track is 100 nm, 500 nm, 1 μm, 2 μm, 5 μm, 10 μm, 50 μm, 100 μm, 300 μm, 500 μm, 1 mm, 2 mm, 5 mm, 1 cm greater than the average thickness of the device, or within the range between any two of these values.

[0527] The device, kit, system or method according to any one of the above embodiments, wherein the average gap size of the slot is 50 μm to 300 μm greater than the average thickness of the device.

[0528] The device, kit, system or method according to any one of the above embodiments, wherein the receiving region of the socket slot is 1 mm larger than the area of the device2 (square millimeters) or less, 10mm 2 or less, 25mm 2 or less, 50mm 2 or less, 75mm 2 or less, 1cm 2 (square centimeters) or less, 2cm 2 or less, 3cm 2 or less, 4cm 2 or less, 5cm 2 or less, 10cm 2 or less, 100cm 2 or less, or within a range between any two of the said values.

[0529] The device, kit, system or method according to any one of the above embodiments, wherein the shape of one or two of the plates is the same as the shape of the socket slot.

[0530] The device, kit, system or method according to any one of the above embodiments, wherein the socket slot has the shape of a box with an open surface, having a length of 31mm, a width of 27mm and a height of 2.5mm.

[0531] The device, kit, system or method according to any one of the above embodiments, wherein the QMAX device is at most only partially located within the socket slot when fully engaged, and the shape of the part of one or two of the plates is the same as the shape of the socket slot.

[0532] Reagent

[0533] The device, kit, system or method according to any one of the above embodiments, wherein the dye for staining WBC is coated on the first plate or the second plate or both.

[0534] The device, kit, system or method according to any one of the above embodiments, wherein the staining dyes for WBC and PLT are coated on the first plate or the second plate or both.

[0535] The device, kit, system or method according to any one of the above embodiments, wherein the dye for staining PLT is coated on the first plate or the second plate or both.

[0536] The device, kit, system or method according to any one of the above embodiments, wherein the reagent is coated in an array by droplet printing.

[0537] The device, kit, system or method according to any one of the above embodiments, wherein the reagent is coated by jetting.

[0538] The device, kit, system or method according to any one of the above embodiments, wherein the reagent is coated by contact printing.

[0539] The device, kit, system or method according to any one of the above embodiments, wherein the reagent is coated by transfer printing.

[0540] The device, kit, system or method according to any one of the above embodiments, wherein the dye for staining RBC is coated on the first plate or the second plate or both.

[0541] The device, kit, system or method according to any one of the above embodiments, wherein the surfactant for separating and rounding RBC is coated on the first plate or the second plate or both.

[0542] The device, kit, system or method according to any one of the above embodiments, wherein the chemical for lysing RBC is coated on the first plate or the second plate or both.

[0543] The device, kit, system or method according to any one of the above embodiments, wherein acridine orange is coated on the first plate or the second plate or both.

[0544] The device, kit, system or method according to any one of the above embodiments, wherein Zwittergent is coated on the first plate or the second plate or both.

[0545] The device, kit, system or method according to any one of the above embodiments, wherein methylene blue and Zwittergent are coated on the first plate or the second plate or both.

[0546] The device, kit, system or method according to any one of the above embodiments, wherein acridine orange and Zwittergent are coated on the first plate or the second plate or both.

[0547] The device, kit, system or method according to any one of the above embodiments, wherein YOYO dye and Zwittergent are coated on the first plate or the second plate or both.

[0548] The device or method according to any one of the above embodiments, wherein the device further comprises a plurality of reagent layers on one or two plates, the plurality of reagent layers including anti-adhesion, cell lysis, cell staining, release time control material layers, and combinations thereof;

[0549] The thickness of each layer coated on the plate is 10 nm, 100 nm, 200 nm, 500 nm, 1 μm, or within a range between any two of these values.

[0550] Among them, the anti-adhesion agent includes ethylenediaminetetraacetic acid (EDTA), disodium ethylenediaminetetraacetate, K2EDTA, K3EDTA, etc.;

[0551] Among them, the cell stain includes Wright's stain (eosin, methylene blue), Giemsa stain (eosin, methylene blue, and azure B), May-grünwald stain, Leishman's stain ("polychrome" methylene blue (i.e., demethylated to azurin) and eosin), erythrosine B stain (erythrosine B), and other fluorescent dyes, including but not limited to acridine orange dye, 3,3'-dihexyloxacarbocyanine (DiOC6), propidium iodide (PI), fluorescein isothiocyanate (FITC), and basic orange 21 (BO21) dye, ethidium bromide, luciferyl sulfate, and diamidinostilbene disulfonic acid derivatives, erythrosine B or trypan blue, Hoechst 33342, trihydrochloride, Trihydrate, DAPI (4,6-diamidino-2-phenylindole, dihydrochloride), YOYO;

[0552] Among them, the cell stain includes Wright's stain (eosin, methylene blue), Giemsa stain (eosin, methylene blue, and azure B), May-grünwald stain, Leishman's stain ("polychrome" methylene blue (i.e., demethylated to azurin) and eosin), erythrosine B stain (erythrosine B), and other fluorescent dyes, including but not limited to acridine orange dye, 3,3'-dihexyloxacarbocyanine (DiOC6), propidium iodide (PI), fluorescein isothiocyanate (FITC), and basic orange 21 (BO21) dye, ethidium bromide, luciferyl sulfate, and diamidinostilbene disulfonic acid derivatives, erythrosine B or trypan blue, Hoechst 33342, trihydrochloride, Trihydrate, DAPI (4,6-diamidino-2-phenylindole, dihydrochloride), YOYO, acid fuchsin, hematoxylin, Hoechst stain, including Hoechst 33258 and Hoechst 33342, methyl green, methylene blue, Nile blue, Nile red, osmium tetroxide, rhodamine, safranin, meosic-AAPV-AMC, CFSE, BCECF / AM, silver nitrate, neutral red, pyronin Y, calcein-AM, dihydroethidium, dimethylxylenecyanol FF, rhodamine 123, 4-methylumbelliferyl palmitate, fast blue B salt, fluorescein, CH dipotassium salt, DAPI dilactate, propidium iodide;

[0553] Among them, the cell lysing agent includes ammonium chloride, sodium bicarbonate, ethylenediaminetetraacetic acid (EDTA), acetic acid, citric acid, other acids and bases, etc.;

[0554] Among them, the release time control material includes albumin, carbomer, carboxymethyl cellulose, carrageenan, chitosan, dextrin, polyethylene glycol, polyvinylpyrrolidone, or polyvinyl alcohol, etc.

[0555] In some embodiments, a chemical with a certain concentration is coated on a plate and dissolved into the blood to achieve a uniform distribution of red blood cells in the device.

[0556] In some embodiments, a chemical with a certain concentration is coated on a plate and dissolved into the blood to lyse red blood cells in the device.

[0557] The coating can be on the first plate or the second plate or both.

[0558] In certain embodiments, the chemicals coated in the device include, but are not limited to, surfactants, Zwittergent, ASB-14, ASB-16, CHAPS, cationic surfactants NN-[tris(hydroxymethyl)methyl]-N-alkyl-N,N-dimethylammonium chloride (lla), llb, llc, lld, CTAC, Tween 20, Tween 40, Tween 60, Tween 80, sodium lauryl sulfate (SLS), ammonium lauryl sulfate, CTAB, sodium lauryl ether sulfate (SLES), sodium laureth sulfate, dioctyl sodium sulfosuccinate, perfluorooctanesulfonate, alkyl-aryl ether phosphate, alkyl ether phosphate, CTAB, cetylpyridinium chloride (CPC), benzalkonium chloride (BAC), benzethonium chloride (BZT), dimethyldioctadecylammonium chloride, dioctadecyldimethylammonium bromide (DODAB), cocamidopropyl hydroxysultaine, cocamidopropyl betaine, narrow range ethoxylates, octaethylene glycol monododecyl ether, pentaethylene glycol monododecyl ether, nonxynols, Triton X-100, polyethoxylated tallow amine, cocoamide monoethanolamine, cocoamide diethanolamine, poloxamer, glycerol monostearate, glycerol monolaurate, sorbitan laurate, sorbitan monostearate, sorbitan tristearate, alkyl polyglucosides, lauryl glucoside, octyl glucoside, lauryldimethylamine oxide, dimethyl sulfoxide, phosphine oxide.

[0559] In some embodiments, the reagents coated in the device that cause red blood cell lysis include, but are not limited to, Pluronic F-127, polyoxyethylene castor oil, Pluronic F-68, Myrj 52, Brij 35, sodium oleate, sodium dodecyl sulfate, Tween 20, Tween 40, Tween 60, Tween 80, SLS, CTAB, CTAC, tamoxifen, saponin, hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, lactic acid, ABS-14, ABS-16, antimalarial drugs (quinine compounds), arsenic, dapsone, metals (chromium / chromate, platinum salts, nickel compounds, copper, lead, cisplatin), nitrite, nitrofurantoin, penicillin, phenazopyridine (marolfen), rho immunoglobulin, ribavirin, sulfonamides, sulfones.

[0560] In some embodiments, the anticoagulants coated in the device include, but are not limited to, EDTA, such as dipotassium ethylenediaminetetraacetate (K2EDTA), tripotassium ethylenediaminetetraacetate (K3EDTA), coumarin (vitamin K antagonist), warfarin (coumadin), acenocoumarol, phenprocoumon, atomoxetin, phenindione, heparin, fondaparinux sodium, and idraparinux, dabigatran etexilate, rivaroxaban, apixaban, edoxaban, betrixaban, NOAC, hirudin, recombinant hirudin, bivalirudin, agratroban, dabigatran etexilate, batroxobin, glossiphoniin, vitamin E, sodium citrate, citrate dextrose, oxalates such as fluoride oxalate, deltaparin, desirudin, enoxaparin.

[0561] In some embodiments, in order to achieve a uniform distribution of red blood cells in the device, Zwittergent is coated on the plate at a preferred areal concentration of 3 ng / mm 2 , 5 ng / mm 2 , 8 ng / mm 2 , 12 ng / mm 2 , 15 ng / mm 2 , 25 ng / mm 2 , 35 ng / mm 2 , 50 ng / mm 2 , 80 ng / mm 2 , or within a range between any two of these values.

[0562] In some embodiments, in order to lyse red blood cells in the device, Zwittergent is coated on the plate at a preferred areal concentration of 100 ng / mm 2 , 120 ng / mm 2 , 150 ng / mm 2 , 180 ng / mm 2 , 200 ng / mm 2 , 300 ng / mm 2 , 400 ng / mm 2 , 500 ng / mm 2 , 800 ng / mm 2 , 1000 ng / mm 2 , or within a range between any two of these values.

[0563] In some embodiments, to achieve a uniform distribution of red blood cells in the device, Zwittergent is coated on the plate at a preferred final concentration in the blood of 0.05 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.5 mg / mL, 0.6 mg / mL, 1.0 mg / mL, 2 mg / mL, or within a range between any two of these values.

[0564] In one embodiment, to lyse red blood cells in the device, Zwittergent is coated on the plate at a preferred final concentration in the blood of 2 mg / mL, 3 mg / mL, 4 mg / mL, 5 mg / mL, 6 mg / mL, 7 mg / mL, 9 mg / mL, 10 mg / mL, 15 mg / mL, 25 mg / mL, 50 mg / mL, or within a range between any two of these values.

[0565] In some embodiments, to achieve a uniform distribution of red blood cells in the device, Zwittergent is coated on the plate at 3 ng / mm 2 , 5 ng / mm 2 , 8 ng / mm 2 , 12 ng / mm 2 , 15 ng / mm 2 , 25 ng / mm 2 , 35 ng / mm 2 , 50 ng / mm 2 , 80 ng / mm 2 , 100 ng / mm 2 , or at a preferred areal concentration within a range between any two of these values.

[0566] In some embodiments, to lyse red blood cells in the device, Zwittergent is coated on the plate at 100 ng / mm 2 , 120 ng / mm 2 , 150 ng / mm 2 , 180 ng / mm 2 , 200 ng / mm 2 , 300 ng / mm 2 , 400 ng / mm 2 , 500 ng / mm 2 , 800 ng / mm 2 , 1000 ng / mm 2 , or at a preferred areal concentration within a range between any two of these values.

[0567] In some embodiments, to achieve a uniform distribution of red blood cells in the device, Zwittergent is coated on the plate at a preferred final concentration in blood of 0.05 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.5 mg / mL, 0.6 mg / mL, 1.0 mg / mL, 2 mg / mL, or within a range between any two of these values.

[0568] In a certain embodiment, to lyse red blood cells in the device, Zwittergent is coated on the plate at a preferred final concentration in blood of 2 mg / mL, 3 mg / mL, 4 mg / mL, 5 mg / mL, 6 mg / mL, 7 mg / mL, 9 mg / mL, 10 mg / mL, 15 mg / mL, 25 mg / mL, 50 mg / mL, or within a range between any two of these values.

[0569] The device, kit, system or method according to any one of the above embodiments, wherein acridine orange is coated on the plate at an area concentration of 0.5 ng / mm 2 , 1 ng / mm 2 , 2 ng / mm 2 , 3 ng / mm 2 , 5 ng / mm 2 , 8 ng / mm 2 , 10 ng / mm 2 , 15 ng / mm 2 , 20 ng / mm 2 , 30 ng / mm 2 , or within a range between any two of these values.

[0570] The device, kit, system or method according to any one of the above embodiments, wherein acridine orange is coated on the plate at an area concentration of 3 to 10 ng / mm 2 , and Zwittergent is coated on the plate at an area concentration of 3 to 10 ng / mm 2 .

[0571] The device, kit, system or method according to any one of the above embodiments, wherein acridine orange is coated on the plate at an area concentration of 5 to 20 ng / mm 2 , and Zwittergent is coated on the plate at an area concentration of 10 to 30 ng / mm 2 .

[0572] Material

[0573] The device, kit, system or method according to any one of the above embodiments, wherein the materials of the first plate and the second plate are poly(methyl methacrylate), polystyrene, polycarbonate, polyethylene terephthalate, polyamide, polyester, polyethylene, polypropylene, polyurethane, polyvinyl chloride, acrylonitrile butadiene styrene, polyepoxide, polytetrafluoroethylene, phenolic, furan, silicone, polylactic acid, polyimide, etc.

[0574] The device, kit, system or method according to any one of the above embodiments, wherein the surfaces of the first plate and the second plate are silicon oxide or silicon nitride.

[0575] The device, kit, system or method according to any one of the above embodiments, wherein the surfaces of the first plate and the second plate are treated to be hydrophilic.

[0576] The device, kit, system or method according to any one of the above embodiments, wherein the surfaces of the first plate and the second plate are treated to not bind cells.

[0577] Transfer pipette

[0578] The device, kit, system or method according to any one of the above embodiments, wherein blood is directly transferred from a finger to the QMAX card.

[0579] The device, kit, system or method according to any one of the above embodiments, wherein blood is transferred from the body surface to the QMAX card using a transfer pipette, and the transfer pipette has a volume of 2 μL, 3 μL, 5 μL, 8 μL, 10 μL, 15 μL or within the range between any two of these values.

[0580] Landing mark

[0581] The device, kit, system or method according to any one of the above embodiments, wherein the landing mark of the blood droplet is on the outer surface of the first plate or the second plate.

[0582] The device, kit, system or method according to any one of the above embodiments, wherein the landing mark of the blood droplet is outside the field of view of the image.

[0583] The device, kit, system or method according to any one of the above embodiments, wherein the landing mark of the blood droplet is close to the center of the card.

[0584] The device, kit, system or method according to any one of the above embodiments, wherein the landing mark of the blood droplet is a small dot or a small cross.

[0585] Filling mark

[0586] The device, kit, system, or method according to any one of the above embodiments, wherein the filling mark for notifying the user of the minimum volume and coverage area of the sample in the device is on the outer surface of the first plate or the second plate.

[0587] The device, kit, system, or method according to any one of the above embodiments, wherein the filling mark for notifying the user of the minimum volume and coverage area of the sample in the device is outside the field of view of the image.

[0588] The device, kit, system, or method according to any one of the above embodiments, wherein the filling mark for notifying the user of the minimum volume and coverage area of the sample in the device is near the center of the card.

[0589] The device, kit, system, or method according to any one of the above embodiments, wherein the filling mark for notifying the user of the minimum volume and coverage area of the sample in the device is circular or rectangular.

[0590] Example QMAX device measures complete blood count and is compared with a commercial machine

[0591] Figure 7 An example device and method for measuring a complete blood count (CBC) using a QMAX device are shown. The device can measure all CBC parameters without dilution. Preliminary tests show that the measurement results of this device are accurate compared with commercial machines.

[0592] Figure 7 Pictures of (a) the QMAX device and (b) the QMAX device and adapter on a smart phone are shown.

[0593] The device is made of PMMA material. The device can be made of polystyrene, PMMA, PC, COC, COP, or other plastic materials.

[0594] In this example, the thickness of plate 1 used is from 950 μm to 1050 μm. Plate 1 has a preferred thickness range of 200 μm to 1500 μm.

[0595] In this example, the thickness of plate 2 used is from 170 μm to 180 μm. Plate 2 has a preferred thickness range of 50 μm to 250 μm.

[0596] A device for measuring RBC and PLT in the experiment has a column height of 5 μm, an inter-column distance of 90 μm, and a column size of 20 μm. The column can have a column height from 2 μm to 6 μm, an inter-column distance from 50 μm to 200 μm, and a column size from 5 μm to 40 μm.

[0597] An apparatus for measuring HgB and WBC in an experiment has a column height of 30 μm, a column pitch of 80 μm, and a column size of 30 μm. The column can have a column height of 20 μm to 50 μm, a column pitch of 50 μm to 200 μm, and a column size of 10 μm to 50 μm.

[0598] On plate 1, acridine orange dye for staining WBC and PLT and Zwittergent for partitioning RBC are coated.

[0599] Acridine orange is coated on the plate at an area concentration of 10 - 80 ng / mm 2 and Zwittergent is coated on the plate at an area concentration of 20 - 130 ng / mm 2 of the area.

[0600] In some other examples, a staining reagent is coated on one or both of the plates. A cell separation reagent is coated on one or both of the plates. The cell lysis reagent is coated on one or both of the plates.

[0601] When measuring and analyzing a whole blood sample using such an apparatus, the following steps are included:

[0602] (a) Obtain a whole blood sample (which can refer to freshly drawn blood or K 2 EDTA venous whole blood) and the apparatus;

[0603] (b) When the plate is configured in an open structure, deposit the sample on one or both of the plates,

[0604] (c) After (b), force the two plates to form a closed structure; and

[0605] (d) When the plate is in the closed structure, shine light on the apparatus and capture an image of the sample in the apparatus; and

[0606] (e) Analyze the image to analyze the whole blood count in the apparatus.

[0607] Figure 8 Shows (a) a bright - field image of HgB in the apparatus at a wavelength of about 520 nm, (b) a bright - field image of WBC in the apparatus at an excitation of about 490 nm and an emission of over 500 nm, (c) a bright - field image of RBC in the apparatus, (d) a bright - field image of WBC and PLT in the apparatus at an excitation of about 490 nm and an emission of over 500 nm, where the interior of the whole blood is photographed by an optical system based on an iPhone.

[0608] In (a), the red blood cells with a column height of 30 μm are multi - layer, which is beneficial for the determination of HgB. In (c), the red blood cells with a column height of 5 μm become monolayer and can be counted in the magnified image.

[0609] White blood cells and platelets were stained with AO dye, and the fluorescence images were bright spots. White blood cells in the 5 μm and 30 μm spacing devices became monolayer and were countable in the magnified images. Platelets in the 5 μm spacing device were monolayer and were countable in the magnified images.

[0610] Whole blood samples (venous blood in K2EDTA tubes) from 50 - 100 patients were measured using the QMAX device and compared with a commercial hematology counter such as the Horiba Pentra 60C. 9 μL of whole blood was dropped onto Plate 2 and squeezed through Plate 1. Then the card was read by the smartphone-based optical system as Figure 8 shown. Cells were counted using local software with OpenCV and machine learning algorithms.

[0611] Figure 9 Exemplary HgB, WBC, RBC, PLT analysis results of whole blood samples using the QMAX device are shown and compared with a commercial hematology counter such as the Horiba Pentra 60C. The results show that the device and method have better accuracy compared to commercial machines.

[0612] Specifically, compared with the Horiba Pentra 60C, in the measurement range of 7 g / dL to 20 g / dL, the HgB readings of the commercial machine had R2 = 98.5%, in the measurement range of 0.4×10 3 / μL to 0.4×10 3 / μL, the WBC readings of the commercial machine had R2 = 99.3%, in the measurement range of 2.3×10 6 / μL to 5.2×10 6 / μL, the RBC readings of the commercial machine had R2 = 98.2%, and in the measurement range of 21×10 3 / μL to 581×10 3 / μL, the WBC readings had R2 = 93%.

[0613] Example 2 Measurement of WBC and WBC differentiation using the QMAX device

[0614] Figure 10 An example result of measuring WBC and WBC differentiation using the QMAX device is shown. This device can measure all WBCs and 3 subtypes (granulocytes, monocytes, lymphocytes) without dilution. Preliminary tests show that the measurement results of this device are accurate compared to commercial machines.

[0615] Figure 10Shows (a) a fluorescence photograph of WBCs in a QMAX device taken by a smartphone optical system, and (b) a summary of the statistical count of WBCs in the device, plotting the comparison of WBC count, green channel intensity, and red channel intensity for each WBC.

[0616] This device is the same as the WBC device in Example 1.

[0617] The acridine orange dye used to stain WBCs is coated on Plate 1. When AO binds to DNA, AO intercalates into DNA as a monomer, producing strong green fluorescence under blue excitation. When it binds to RNA and proteins, it forms an electrostatic complex in polymer form, producing red fluorescence under blue excitation. Since the DNA / RNA ratios of the three subtypes (granulocytes, monocytes, lymphocytes) are different, the differentiation of WBCs can be achieved by analyzing the green and red fluorescence ratios of each WBC.

[0618] Acridine orange is coated on the plate at an area concentration of 10 - 80 ng / mm 2 and Zwittergent is coated on the plate at an area concentration of 20 - 130 ng / mm 2 of the area concentration.

[0619] When measuring and analyzing a whole blood sample using such a device, it includes the following steps:

[0620] (a) Obtain a whole blood sample (which can be fingertip blood or K 2 EDTA venous whole blood) and the device;

[0621] (b) When the plate is configured in an open structure, deposit the sample on one or both of the plates,

[0622] (c) After (b), force the two plates to form a closed structure; and

[0623] (d) When the plate is in the closed structure, shine light on the device and capture an image of the sample in the device; and

[0624] (e) Analyze the image to analyze the whole blood count in the device.

[0625] The wavelength of the excitation irradiation is 450 - 480 nm, and the emission is a long - pass with a cut - off wavelength of around 520 nm. The green (550 nm) and red (650 nm) fluorescence of each WBC can be observed from the camera.

[0626] According to the results, white blood cells stained with AO dye are colored dots on the fluorescence image, with distinct three colors (green, yellow, and red), as Figure 10(a), which corresponds to lymphocytes (more DNA), monocytes (balanced DNA, RNA), and granulocytes (more RNA). Machine learning and software are used to analyze the color of each WBC and classify it into 3 clusters, as shown in Figure 10 (b).

[0627] iMOST HgB+WBC+WBC differentiation QMAX card example

[0628] In a preferred embodiment, the spacing between the plates and / or the sample thickness is about 30 μm.

[0629] In a preferred embodiment, the spacing between the plates and / or the sample thickness is between 20 μm and 40 μm.

[0630] The spacer is a rectangular shape with rounded corners.

[0631] The lateral dimension of the spacer is about 30 μm × 40 μm.

[0632] The lateral dimension of the spacer is 10 μm to 40 μm.

[0633] The diameter of the rounded corners of the spacer is 10 μm.

[0634] The spacer is in a rectangular dot matrix array.

[0635] The spacer pitch of the spacer is about 80 μm.

[0636] The spacer pitch of the spacer is 70 μm to 150 μm.

[0637] The length of one plate of the Q card is 27 mm, and the width of the plate is 22 mm.

[0638] The length of one plate of the Q card is 32 mm, and the width is 24 mm.

[0639] The area of one plate is about 600 mm 2 , and the area of the other plate is about 750 mm 2 .

[0640] The thickness of one plate of the Q card is about 175 μm.

[0641] The thickness of one plate of the Q card is about 1 mm.

[0642] The notch area on the QMAX card is in the range of 10 to 30 mm 2 .

[0643] The notch is semi-circular with a diameter of 3 - 6 mm.

[0644] The notch has a width of 3 mm and a length of 6 mm.

[0645] The width of the hinge joint is about 6 mm.

[0646] The length of the hinge joint is about 20 mm.

[0647] The hinge has a thickness of about 70 μm.

[0648] The reagent is coated in an array by droplet printing.

[0649] The reagent is coated by spraying.

[0650] Acridine orange or other staining reagents are coated onto the first plate or the second plate or both.

[0651] Zwittergent or other detergents are coated onto the first plate or the second plate or both.

[0652] Acridine orange is coated on the plate at an area concentration of 10 - 60 ng / mm 2 and Zwittergent is coated on the plate at an area concentration of 20 - 130 ng / mm 2 on the plate.

[0653] The materials of the first plate and the second plate are poly(methyl methacrylate).

[0654] The landing mark of the blood droplet is on the outer surface of the first plate or the second plate.

[0655] The landing mark of the blood droplet is a small dot or a small cross.

[0656] The landing mark of the blood droplet is outside the field of view of the image.

[0657] The landing mark of the blood droplet is close to the center of the card.

[0658] At least one of the plates is transparent.

[0659] iMOST RBC+PLT QMAX card example

[0660] Same as the above 1, except that:

[0661] In a preferred embodiment, the spacing between the plates and / or the sample thickness is about 5 μm.

[0662] In a preferred embodiment, the spacing between the plates and / or the sample thickness is between 2 μm and 7 μm.

[0663] The lateral dimension of the spacer is about 30 μm to 40 μm.

[0664] The lateral dimension of the spacer is 5 μm to 40 μm.

[0665] Acridine orange or other staining reagents are coated onto the first plate or the second plate or both.

[0666] Zwittergent or other detergents are coated onto the first plate or the second plate or both.

[0667] Acridine orange is coated onto the plate at an area concentration of 10 - 60 ng / mm 2 and Zwittergent is coated onto the plate at an area concentration of 20 - 130 ng / mm 2 of the area.

Claims

1. An apparatus for analyzing white blood cells in a blood sample, comprising: a first plate, a second plate, and a spacer, wherein: i. The two plates are movable relative to each other into different configurations; ii. One or both of the plates are flexible; iii. Each of the two plates includes an inner surface having a sample contact area for contacting the blood sample; iv. One or both of the two plates include the spacer permanently fixed to the sample contact area of the corresponding plate; v. The spacer has: (a) a predetermined uniform height having a value selected in the range of 2 µm to 50 µm, (b) the shape of a column having a uniform cross-section and a flat top surface; (c) a width-to-height ratio equal to or greater than 1; (d) a predetermined fixed, non-random spacer spacing in the range of 10 µm to 200 µm; (e) a fill factor equal to 1% or greater, where the fill factor is the ratio of the contact area of the spacer fixed on the plate to the total area of the plate; and (f) the product of the fill factor and the Young's modulus of the spacer is 2 MPa or greater; wherein one of the configurations is an open configuration in which the two plates are partially or fully separated, the spacing between the two plates is not adjusted by the spacer, and the sample is deposited on one or both of the two plates; and wherein the other of the configurations is a closed configuration configured after the sample is deposited in the open configuration; and in the closed configuration, at least a portion of the sample is compressed by the two plates into a layer of very uniform thickness and is stagnant relative to the two plates, wherein the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the two plates and the spacer.

2. An apparatus for analyzing white blood cells in a blood sample, comprising: a first plate, a second plate, a spacer, and an adapter, wherein: i. The two plates are movable relative to each other into different configurations; ii. One or both of the plates are flexible; iii. Each of the two plates includes an inner surface having a sample contact area for contacting the fluid sample; iv. One or both of the two plates include the spacer permanently fixed to the sample contact area of the corresponding plate; v. The spacer has: (a) a predetermined uniform height having a value selected in the range of 2 µm to 50 µm, (b) the shape of a column having a uniform cross-section and a flat top surface; (c) a width-to-height ratio equal to or greater than 1; (e) a predetermined fixed, non-random spacer spacing in the range of 10 µm to 200 µm; (e) a fill factor equal to 1% or greater, where the fill factor is the ratio of the contact area of the spacer fixed on the plate to the total area of the plate; and (f) the product of the fill factor and the Young's modulus of the spacer is 2 MPa or greater; ​ ​ ​ vi. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone having a camera, (c) a slot in the housing that allows (1) a plate in a closed configuration to slide into the slot, and (2) when the plate is in the slot, at least a portion of the sample area is less than 2 cm from the outer surface of the camera, and (d) an optical system in the housing configured to image at least a portion of the sample contact area by the camera; One of the configurations is an open configuration in which the two plates are partially or fully separated, the spacing between the two plates is not adjusted by the spacer, and the sample is deposited on one or both of the plates; and The other of the configurations is a closed configuration configured after the sample is deposited in the open configuration; And in the closed configuration, at least a portion of the sample is compressed into a layer with a very uniform thickness and is stagnant relative to the two plates, wherein the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the two plates and the spacer.

3. A device for analyzing white blood cells in a blood sample, comprising: a first plate, a second plate, a spacer, and an adapter, wherein: i. The two plates are movable relative to each other into different configurations; ii. One or both of the plates are flexible; iii. Each of the two plates includes an inner surface having a sample contact area for contacting a fluid sample; iv. One or both of the two plates include the spacer permanently fixed on the sample contact area of the corresponding plate spacer; v. The spacer has: (a) a predetermined uniform height having a value selected in the range of 10 µm to 50 µm, (b) the shape of a column having a uniform cross-section and a flat top surface; (c) the ratio of the width to the height is equal to or greater than 1; (f) a predetermined fixed, non-random spacer spacing in the range of 10 µm to 200 µm; (e) a packing factor equal to 3% or greater, where the packing factor is the ratio of the contact area of the spacer fixed on the plate to the total area of the plate; and (f) the product of the packing factor and the Young's modulus of the spacer is 2 MPa or greater; vi. One or both of the two plates include a reagent coated on the sample contact area of the corresponding plate; vii. The reagent has at least one of the following: (a) a component that stains WBC; (b) a component that makes RBCs evenly distributed; (c) a component that lyses RBCs; viii. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone having a camera, (c) a slot in the housing that allows (1) a plate in a closed configuration to slide into the slot, and (2) when the two plates are in the slot, at least a portion of the sample area is less than 2 cm from the outer surface of the camera, and (d) an optical system in the housing configured to image at least a portion of the sample contact area by the camera; 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 a spacer, and the sample is deposited on one or both of the plates; and The other of the configurations is a closed configuration that is configured after the sample is deposited in the open configuration; And in the closed configuration, at least a portion of the sample is compressed into a layer of very uniform thickness and is stationary relative to the plates, wherein the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the plates and the spacer.

4. A method for analyzing an analyte in a liquid sample, comprising: (a) obtaining the liquid sample; (b) obtaining the device according to any one of claims 1 to 3; (c) depositing the liquid sample on one or both of the two plates when the two plates are configured in the open configuration; (d) after (c), forcing the two plates to form a closed configuration and compressing at least a portion of the sample into a layer of uniform thickness; (e) capturing an image of the sample in the layer of uniform thickness using a camera, wherein the image shows the analyte; and (f) analyzing the image to enumerate the analyte in the image; (g) calculating the concentration of the analyte in the sample based on the uniform thickness, the field of view (FoV) of the camera, the analyte enumeration, and a predetermined correction factor; wherein the field of view is the range of the field in which the camera captures the image; wherein the correction factor is determined by an error counting ratio that depends on the field of view, the uniform thickness, and the nature of the analyte.

5. A method for analyzing white blood cells in a blood sample, comprising: (a) obtaining a blood sample; (b) obtaining the device according to any one of claims 1 to 3; wherein (e) capturing an image of the sample in the layer of uniform thickness when the two plates are in the closed configuration; and (g) analyzing the image to determine the concentration of white blood cells in the sample.

6. A method for performing white blood cell and subtype counting using a single device, comprising: (a) obtaining a blood sample; (b) obtaining the device according to any one of claims 1 to 3, wherein the spacer height is 5 µm to 40 µm, (c) depositing the blood sample on one or both of the two plates when the two plates are configured in the open configuration on, (d) after (c), forcing the two plates to form a closed configuration; (e) capturing an image of the sample in the layer of uniform thickness when the two plates are in the closed configuration; and (f)Analyze the image to determine the respective quantities of white blood cells, neutrophils, lymphocytes, monocytes, eosinophils, and basophils by counting the number of cells in the image and analyzing the fluorescence color and shape of each white blood cell.

7. The method according to claim 4, wherein, the method for measuring an analyte in a liquid sample comprises: (a)Insert a sample card composed of the first plate and the second plate into an adapter configured to be attached to a device comprising a light source and a camera; (b)Irradiate the sample with light from the light source, wherein: i. The light is filtered by an optical filter of the adapter device to form a first light beam, a part of the first light beam irradiates the edge of the sample card and travels in the sample card to irradiate the sample; ii. A part of the first light beam is deflected by a mirror of the adapter device to form a second light beam that irradiates the sample in an inclined angle in the reverse direction; and iii. The remaining part of the first light beam with a divergence angle is absorbed by an absorber of the adapter device.

8. The method according to claim 7, wherein, further comprising: (e)Capture an image of the sample in the uniform thickness layer using a camera; (f)Analyze the image to enumerate the analyte in the image; and (g)Calculate the concentration of the analyte in the sample based on the uniform thickness, the field of view of the camera, the number of analytes, and a predetermined correction factor; wherein the field of view is the range of the field where the camera captures the image; wherein the correction factor is determined by an error counting ratio, and the error counting ratio depends on the field of view, the uniform thickness, and the nature of the analyte.

9. The method according to claim 5 or 6, wherein, the blood sample is undiluted.

10. The method according to claim 6, wherein, The staining and shape of white blood cells provide fluorescence color, structure, and size differentiation of white blood cells and their subtypes, as well as white blood cell differentiation.

11. The method according to claim 10, wherein, The fluorescence color is used for WBC counting and differentiation.

12. The method according to claim 10, wherein, The size of white blood cells is used for WBC counting and differentiation.

13. The method according to claim 10, wherein, The structure of white blood cells is used for WBC counting and differentiation.

14. The method according to claim 10, wherein, Both the fluorescence color and structure of white blood cells are used for WBC counting and differentiation.

15. The method according to claim 5 or 6, wherein, The color of white blood cells is distinguished by the red, green, and blue channels of the image.

16. The method according to claim 5 or 6, wherein the color of white blood cells is distinguished by filters at different wavelengths in front of the camera.

17. The method according to claim 5 or 6, wherein, The color of white blood cells is distinguished by filters at different wavelengths in front of the light source.

18. The method according to claim 5 or 6, wherein, The color of the white blood cells comes from the staining of WBC by acridine orange dye.

19. The method according to claim 5 or 6, wherein, the color of the white blood cells comes from the staining of WBC by different chemicals.

20. The method according to claim 5 or 6, wherein, the color of the white blood cells comes from the staining of WBC by methylene blue.

21. The device according to any one of claims 1 to 3, wherein a staining agent is coated on one or both of the two plates.

22. The device according to any one of claims 1 to 3, wherein a cell separation reagent is coated on one or both of the two plates.

23. The device according to any one of claims 1 to 3, wherein a cell lysis reagent is coated on one or both of the two plates.

24. The method according to claim 5 or 6, wherein the shape of the white blood cells is analyzed by machine learning.

25. The method according to claim 5 or 6, wherein the color and shape of the white blood cells are analyzed by machine learning.

26. The device according to any one of claims 1 to 3, wherein, Acridine orange is coated on the plate at an area concentration of 1 to 20 ng / mm 2 , and Zwittergent is coated on the plate at an area concentration of 1 to 30 ng / mm 2 .

27. The device according to any one of claims 1 to 3, wherein, one or both of the plates further comprise a plurality of reagent layers, and the plurality of reagent layers include: anti-adhesion, cell lysis, cell staining or release time control material layers.

28. The device according to claim 2 or 3, wherein, the adapter comprises: (a)An attachment member configured to attach the adapter to a device comprising a light source and a camera; (b)A card slot configured to accommodate a sample card formed by the first plate and the second plate, the sample card containing a liquid sample compressed into a uniformly thick layer, wherein when the sample card is inserted into the card slot, the sample is under the field of view of the camera and the light source; (c)An optical filter configured to filter the light from the light source to form a first light beam, wherein a part of the first light beam irradiates the edge of the sample card and travels in the sample card to irradiate the sample; (d)A mirror configured to deflect a part of the first light beam to form a second light beam that irradiates the sample in an inclined angle in the reverse direction; (e)An absorber configured to absorb the remaining part of the first light beam having a divergence angle.

29. The device according to claim 28, wherein, the adapter device further comprises a housing member.

30. The device according to claim 29, wherein, the adapter device further comprises a lever that can be inserted into or pulled out of the housing member.

31. The device according to claim 30, wherein the mirror and the absorber are mounted on the lever.

32. The device according to claim 2 or 3, wherein the adapter comprises a card slot having a safety opening that allows the sample card to be inserted and prevents ambient light from entering the card slot.

33. The method according to claim 8, wherein, The distance from the sample to the camera lens is in the range of 2 mm to 5 mm.

34. The method according to claim 8, wherein, the distance from the sample to the camera lens is in the range of 4 mm to 7 mm.

35. The method according to claim 8, wherein, the distance from the sample to the camera lens is in the range of 6 mm to 9 mm.

36. The method according to claim 8, wherein, the distance from the sample to the camera lens is in the range of 8 mm to 11 mm.

37. The method according to claim 8, wherein, the distance from the sample to the camera lens is in the range of 10 mm to 13 mm.

38. The method according to claim 8, wherein, the distance from the sample to the camera lens is in the range of 12 mm to 15 mm.

39. The method according to any one of claims 4 to 6, further comprising a plurality of reagent layers on one or two plates, the reagent layers comprising: an anti - adhesion reagent, a cell lysis reagent, a cell separation reagent, a cell staining reagent or a release time - control material.

40. The method according to claim 39, wherein, each reagent layer coated on the plate has a thickness of 10 nm, 100 nm, 200 nm, 500 nm or 1 µm.

41. The method according to claim 39, wherein the wettability between the plate and the coating buffer is used to coat the reagent layer.

42. The method according to claim 41, wherein the reagent for coating is dissolved in an organic solvent, the organic solvent comprising: alcohols, ether, hexane, tetrachloroethane, toluene and xylene.

43. The method according to claim 39, wherein, the cell staining reagent comprises: Wright's stain, Giemsa stain, May - Grünwald stain, Leishman's stain, Erythrosin B stain, acridine orange dye, 3,3 - dihexyloxacarbocyanine, propidium iodide, fluorescein isothiocyanate and basic orange 21 dye, ethidium bromide, Brilliant Sulfaflavine and diamidostilbene disulfonic acid derivatives, trypan blue, Hoechst 33342, trihydrochloride, trihydrate, or DAPI.

44. The method according to claim 39, wherein, the cell separation reagent comprises a surfactant; the surfactant includes: Zwittergent, CHAPS, CTAC, Tween 20, Tween 40, Tween 60, Tween 80, SLS, or CTAB.

45. The method according to claim 39, wherein, the cell lysis reagent comprises: ammonium chloride, sodium bicarbonate, ethylenediaminetetraacetic acid (EDTA), acetic acid, or citric acid.

46. The method according to claim 5 or 6, wherein, white blood cells are stained with propidium iodide (PI), fluorescein isothiocyanate (FITC) and basic orange (BOM); PI is a red nucleic acid dye that stains DNA and shows red fluorescence; FITC stains cytoplasmic proteins and shows green fluorescence; BO21 is a green nucleic acid dye that stains DNA and shows green fluorescence; Lymphocytes, monocytes, neutrophils, and eosinophils are distinguished by a combination of three fluorescent dyes.

47. The method according to claim 39, wherein, the anti-adhesion agent comprises: ethylenediaminetetraacetic acid (EDTA), disodium ethylenediaminetetraacetate, K2EDTA, or K3EDTA.

48. The method according to claim 39, wherein, the release time control material comprises albumin, carbomer, carboxymethyl cellulose, carrageenan, chitosan, dextrin, polyethylene glycol, polyvinylpyrrolidone, or polyvinyl alcohol.

49. The method according to claim 5 or 6, wherein, RBCs, platelets, or both in the sample are lysed before detecting and / or measuring WBCs.

50. The method according to claim 5 or 6, wherein WBCs, platelets, or both are lysed in the sample before detecting RBCs.

51. The method according to claim 5 or 6, wherein RBCs, WBCs, or both are lysed in the sample before detecting PLTs.

52. The device according to claim 2 or 3, further comprising: (a)An attachment member configured to attach the adapter to a device comprising a light source and a camera; (b)A card slot configured to receive a sample card formed by the first plate and the second plate, the sample card containing a liquid sample compressed into a uniformly thick layer, wherein when the sample card is inserted into the card slot, the sample is within the field of view of the camera and the light source; (c)An optical filter configured to filter light from the light source to form a first light beam, wherein a portion of the first light beam irradiates an edge of the sample card and travels through the sample card to irradiate the sample; (d)A mirror configured to deflect a portion of the first light beam to form a second light beam that irradiates the sample at an inclined angle in the reverse direction; An absorber configured to absorb the remaining portion of the first light beam having a divergence angle.

53. A method for analyzing an analyte in a liquid sample, comprising: (a)Obtaining the liquid sample and the device according to claim 52; (b)Compressing at least a portion of the sample into a uniformly thick layer using the sample card, (c)Inserting the sample card into an adapter device configured to attach to a device comprising a light source and a camera; (d)Irradiating the sample with light from the light source, wherein: i. The light is filtered by the optical filter of the adapter device to form a first light beam, a portion of the first light beam irradiates an edge of the sample card and travels through the sample card to irradiate the sample; ii. A portion of the first light beam is deflected by the mirror of the adapter device to form a second light beam that irradiates the sample at an inclined angle in the reverse direction; and ​ ​ ​ iii. The remaining part of the first light beam with a divergence angle is absorbed by the absorber of the adapter device.

54. A device for analyzing white blood cells in a blood sample, comprising: a first plate, a second plate, a spacer, and an adapter, wherein: i. The two plates are movable relative to each other into different configurations; ii. One or both of the plates are flexible; iii. Each of the two plates includes an inner surface having a sample contact area for contacting a fluid sample; iv. One or both of the two plates include the spacer permanently fixed on the sample contact area of the corresponding plate; viii. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone having a camera, (c) a slot in the housing that allows (1) the plates in a closed configuration to slide into the slot, and (2) when the two plates are in the slot, at least a part of the sample area is less than 2 cm from the outer surface of the camera, and (d) an optical system in the housing configured to image at least a part of the sample contact area by the camera; wherein one of the configurations is an open configuration in which the two plates are partially or fully separated, the spacing between the two plates is not adjusted by the spacer, and the sample is deposited on one or both of the plates; and wherein the other of the configurations is a closed configuration configured after the sample is deposited in the open configuration; and in the closed configuration, at least a part of the sample is compressed by the two plates into a layer with a very uniform thickness and is stagnant relative to the two plates, wherein the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the two plates and the spacer.

55. The device according to claim 54, wherein the spacer height, the spacing between the two plates, and / or the sample thickness is 30 µm.

56. The device according to any one of claims 1 to 3, 54, wherein, ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ The distance between the two plates and / or the thickness of the sample is 10 µm to 50 µm.

57. The device according to any one of claims 1 to 3, 54, wherein, the spacer is a rounded rectangle.

58. The device according to any one of claims 1 to 3, 54, wherein, the lateral dimension of the spacer is 30 µm to 40 µm.

59. The device according to any one of claims 1 to 3, 54, wherein the lateral dimension of the spacer is 10 µm to 50 µm.

60. The device according to any one of claims 1 to 3, 54, wherein, the rounded diameter of the spacer is 5 µm to 15 µm.

61. The device according to any one of claims 1 to 3, 54, wherein, the spacer is an array of rectangular dots.

62. The device according to any one of claims 1 to 3, 54, wherein, the spacing pitch of the spacer is 80 µm.

63. The device according to any one of claims 1 to 3, 54, wherein, the spacing pitch of the spacer is 70 µm to 150 µm.

64. The device according to any one of claims 1 to 3, 54, wherein the length of one plate is 27 mm and the width of this plate is 22 mm.

65. The device according to any one of claims 1 to 3, 54, wherein the length of one plate is 32 mm and the width of this plate is 24 mm.

66. The device according to any one of claims 1 to 3, 54, wherein the area of one plate is 600 mm 2 , and the area of the other plate is 750 mm 2 .

67. The device according to any one of claims 1 to 3, 54, wherein the thickness of one plate is 175 µm.

68. The device according to any one of claims 1 to 3, 54, wherein the thickness of one plate is 1 mm.

69. The device according to any one of claims 1 to 3 and 54, wherein the one plate has a notch with an area in the range of 10 mm 2 to 30 mm 2 .

70. The device according to claim 69, wherein, the notch is a semi-circular shape with a diameter of 3 mm to 6 mm.

71. The device according to claim 69, wherein, the notch has a width of 3 mm and a length of 6 mm.

72. The device according to any one of claims 1 to 3, 54, the plates are connected by a hinge, and the width of the hinge joint is 6 mm.

73. The device according to claim 72, wherein, the length of the hinge joint is 20 mm.

74. The device according to claim 72, wherein, the hinge has a thickness of 70 µm.

75. The device according to claim 27, wherein, the reagent is coated in an array by droplet printing.

76. The device according to claim 27, wherein, the reagent is coated by spraying.

77. The device according to claim 27, wherein, the reagent is dissolved in more than 70% alcohol for coating.

78. The device according to claim 27, wherein, the reagent coating uses the wetting property between the plate and the reagent solvent.

79. The device according to claim 26, wherein, the acridine orange is coated on the first plate or the second plate or both.

80. The device according to claim 26, wherein, The Zwittergent is coated on the first plate or the second plate or both.

81. The device according to claim 26, wherein, The acridine orange is coated on the plate at an area concentration of 20 to 80 ng / mm 2 and the Zwittergent is coated on the plate at an area concentration of 30 to 120 ng / mm2.

82. The device according to claim 81, wherein, The material at the coating positions of the first plate and the second plate is polymethyl methacrylate.

83. The device according to claim 82, wherein, The landing mark for the blood droplet is on the outer surface of the first plate or the second plate.

84. The device according to claim 83, wherein, The landing mark for the blood droplet is a small dot or a small cross.

85. The device according to claim 83, wherein, The landing mark for the blood droplet is outside the field of view of the image.

86. The device according to claim 83, wherein, The landing mark for the blood droplet is close to the center of the two plates.

87. The device according to claim 54, wherein, A separate paper or plate between the two plates is used to protect the reagent coating on the plates.

88. The device according to claim 54, wherein, A separate paper or plate between the two plates is used to pick up and open the device.

89. The device according to claim 54, wherein, It has a filling mark for notifying the user of the minimum volume and coverage area of the sample in the device.

90. The device according to claim 54, wherein, At least one of the two plates is transparent.

91. The device according to claim 54, wherein, The adapter has a band - pass excitation filter with a transmission band from 450 nm to 480 nm.

92. The device according to claim 54, wherein, The adapter has a long - pass emission filter with a cut - off wavelength between 510 nm and 525 nm.

93. The device according to claim 54, wherein, The adapter has an excitation light irradiation with an oblique angle greater than the collection angle of the imaging optics.

94. The device according to claim 54, wherein, The adapter has a focal length between 2 mm and 6 mm.

95. A device for analyzing red blood cells in a blood sample, comprising: A first plate, a second plate, a spacer, and an adapter, wherein: i. The two plates can be moved relative to each other into different configurations; ii. One or both of the plates are flexible; iii. Each of the two plates contains an inner surface having a sample contact area for contacting the fluid sample; iv. One or both of the two plates contain the spacer permanently fixed on the sample contact area of the corresponding plate; v. The spacer has: (a) A predetermined uniform height, which has a value selected in the range of 10 µm to 150 µm, (b) The shape of a column, which has a uniform cross - section and a flat top surface; (c) The ratio of the width to the height is equal to or greater than 1; (d) A predetermined fixed, non - random spacer spacing in the range of 10 µm to 200 µm (e) A filling factor equal to or greater than 3%, where the filling factor is the ratio of the contact area of the spacer fixed on the plate on the plate to the total area of the plate; and (f) The product of the filling factor and the Young's modulus of the spacer is 2 MPa or greater; vi. One or both of the two plates contain a reagent coated on the sample contact area of the corresponding plate; vii. The reagent has at least one of the following: (a) A component that makes the red blood cells evenly distributed; (b) A component that lyses the red blood cells; (c) A component that dilutes the blood; viii. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone with a camera, (c) a slot in the housing that allows (1) the plate in the closed configuration to slide into the slot, and (2) when the two plates are in the slot, at least a part of the sample area is less than 2 cm from the outer surface of the camera; where one of the configurations is an open configuration, in which the two plates are partially or completely separated, the spacing between the two plates is not adjusted by the spacer, and the sample is deposited on one or both of the two plates; and where the other of the configurations is a closed configuration configured after the sample is deposited in the open configuration; and in the closed configuration, at least a part of the sample is compressed by the two plates into a layer with a very uniform thickness and is stagnant relative to the two plates, where the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the two plates and the spacer.

96. The device according to claim 95, wherein the spacer height, the spacing between the two plates, and / or the sample thickness is 30 µm.

97. The device according to claim 95, wherein, the spacing between the two plates, and / or the sample thickness is 10 µm to 100 µm.

98. The device according to claim 95, wherein, the spacer is a rounded rectangle.

99. The device according to claim 95, wherein, the lateral dimension of the spacer is 30 µm to 40 µm.

100. The device according to claim 95, wherein, the lateral dimension of the spacer is 10 µm to 50 µm.

101. The device according to claim 95, wherein, the rounded corner diameter of the spacer is 5 µm to 15 µm.

102. The device according to claim 95, wherein, the spacer is an array of rectangular dots.

103. The device according to claim 95, wherein, the spacing pitch of the spacer is 80 µm.

104. The device according to claim 95, wherein, the spacing pitch of the spacer is 70 µm to 150 µm.

105. The device according to claim 95, wherein, the length of one plate is 27 mm, and the width of this plate is 22 mm.

106. The device according to claim 95, wherein, ​ The length of a plate is 32 mm and the width of this plate is 24 mm.

107. The device according to claim 95, wherein the area of one plate is 600 mm 2 , and the area of the other plate is 750 mm 2 .

108. The device according to claim 95, wherein the thickness of a plate is 175 µm.

109. The device according to claim 95, wherein the thickness of a plate is 1 mm.

110. The device according to claim 95, wherein, The reagent is dissolved in more than 70% alcohol for coating.

111. The device according to claim 110, wherein, The reagent coating uses the wetting property between the plate and the reagent solvent.

112. The device according to claim 110, wherein, The reagent is coated by spraying.

113. The device according to claim 95, wherein, Zwittergent is coated onto the first plate or the second plate or both.

114. The device according to claim 113, wherein, The Zwittergent is coated on the plate at an area concentration of 20 to 150 ng / mm 2 2.

115. The device according to claim 114, wherein, The material at the coating position of the first plate and the coating position of the second plate is polymethyl methacrylate.

116. The device according to claim 95, wherein, It has a blood drop landing mark, which is on the outer surface of the first plate or the second plate.

117. The device according to claim 116, wherein, The landing mark of the blood drop is a small dot or a small cross.

118. The device according to claim 116, wherein, The landing mark of the blood drop is outside the field of view of the image.

119. The device according to claim 116, wherein, The landing mark of the blood drop is near the center of the two plates.

120. The device according to claim 95, wherein, A separate paper or plate between the two plates is used to protect the reagent coating on the plates.

121. The device according to claim 95, wherein, A separate paper or plate between the two plates is used to pick up and open the device.

122. The device according to claim 95, wherein, It has a filling mark for notifying the user of the minimum volume and coverage area of the sample in the device.

123. The device according to claim 95, wherein, At least one of the two plates is transparent.

124. The device according to claim 95, wherein, The adapter has a band-pass filter with a transmission band from 520 nm to 580 nm.

125. The device according to claim 95, wherein the adapter has a light irradiation angle of 85 degrees to 95 degrees with respect to the surface of the sample.

126. The device according to claim 95, wherein, The adapter has a focal length between 2 mm and 4 mm.

127. A device for analyzing red blood cells in a blood sample, comprising: A first plate, a second plate, a spacer, and an adapter, wherein: i. The two plates can move relative to each other into different configurations; ii. One or both plates are flexible; iii. Each of the two plates includes an inner surface having a sample contact area for contacting a fluid sample; iv. One or both of the two plates include the spacer permanently fixed on the sample contact area of the corresponding plate; v. The spacer has: (a) a predetermined uniform height having a value selected in the range of 1.5 µm to 8 µm; (b) the shape of a column having a uniform cross-section and a flat top surface; (c) a width-to-height ratio equal to or greater than 1; (d) a predetermined fixed, non-random spacer pitch in the range of 10 µm to 200 µm; (e) a fill factor equal to 3% or greater, where the fill factor is the ratio of the contact area of the spacer fixed on the plate to the total area of the plate; and (f) the product of the fill factor and the Young's modulus of the spacer is 2 MPa or greater; vi. One or both of the two plates include a reagent coated on the sample contact area of the corresponding plate; vii. The reagent has at least one of the following: (a) a component that equalizes the distribution of red blood cells; (b) a component that reduces the aggregation of red blood cells; (c) a component that stains red blood cells; (d) a component that dilutes blood; viii. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone having a camera, (c) a slot in the housing that allows (1) the plates in the closed configuration to slide into the slot, and (2) when the two plates are in the slot, at least a portion of the sample area is less than 2 cm from the outer surface of the camera, and (d) an optical system in the housing configured to image at least a portion of the sample contact area by the camera; One of the configurations is an open configuration in which the two plates are partially or fully separated, the spacing between the two plates is not adjusted by the spacer, and the sample is deposited on one or both of the two plates; And the other of the configurations is a closed configuration configured after the sample is deposited in the open configuration; and in the closed configuration, at least a portion of the sample is compressed into a layer with a very uniform thickness and is stagnant relative to the two plates, where the uniform thickness of the layer is limited by the sample contact areas of the two plates and adjusted by the two plates and the spacer.

128. The device according to claim 127, wherein the spacer height, the spacing between the two plates, and / or the sample thickness is 5 µm.

129. The device according to claim 127, wherein, the spacing between the two plates, and / or the sample thickness is 1.5 µm to 8 µm.

130. The device according to claim 127, wherein, the spacer is a rounded rectangle.

131. The device according to claim 127, wherein, the lateral dimension of the spacer is 30 µm to 40 µm.

132. The device according to claim 127, wherein, The lateral dimension of the spacer is 5 µm to 50 µm.

133. The device according to claim 127, wherein, the rounded corner diameter of the spacer is 5 µm to 15 µm.

134. The device according to claim 127, wherein, the spacer is a rectangular dot array.

135. The device according to claim 127, wherein, the spacing between the spacers is 80 µm.

136. The device according to claim 127, wherein, the spacing between the spacers is 70 µm to 150 µm.

137. The device according to claim 127, wherein, the length of one plate is 27 mm and the width of this plate is 22 mm.

138. The device according to claim 127, wherein the length of one plate is 32 mm and the width of this plate is 24 mm.

139. The device according to claim 127, wherein, The area of one board is 600mm 2 , and the area of another board is 750mm 2 .

140. The device according to claim 127, wherein, the thickness of one plate is 175 µm.

141. The device according to claim 127, wherein the thickness of one plate is 1 mm.

142. The device according to claim 127, wherein, the reagent is dissolved in more than 70% alcohol for coating.

143. The device according to claim 142, wherein, the reagent coating uses the wetting property between the plate and the reagent solvent.

144. The device according to claim 142, wherein, the reagent is coated by spraying.

145. The device according to claim 127, wherein, Zwittergent is coated on the first plate or the second plate or both.

146. The device according to claim 145, wherein the Zwittergent is coated on the plate at an areal concentration of 10 ng / mm 2 to 120 ng / mm 2 of the area.

147. The device according to claim 145, wherein, the material at the coating position of the first plate and the coating position of the second plate is polymethyl methacrylate.

148. The device according to claim 145, wherein, it has a landing mark for a blood drop, which is on the outer surface of the first plate or the second plate.

149. The device according to claim 148, wherein, the landing mark of the blood drop is a small dot or a small cross.

150. The device according to claim 148, wherein, the landing mark of the blood drop is outside the field of view of the image.

151. The device according to claim 148, wherein, the landing mark of the blood drop is near the center of the two plates.

152. The device according to claim 127, wherein, a separate paper or plate between the two plates is used to protect the reagent coating on the plates.

153. The device according to claim 127, wherein, a separate paper or plate between the two plates is used to pick up and open the device.

154. The device according to claim 127, wherein, it has a filling mark for notifying the user of the minimum volume and coverage area of the sample in the device.

155. The device according to claim 127, wherein, at least one of the two plates is transparent.

156. The device according to claim 127, wherein, the adapter has a light irradiation angle of 85 degrees to 95 degrees.

157. The device according to claim 127, wherein, the adapter has a focal length between 2 mm and 4 mm.

158. A device for analyzing platelets in a blood sample, comprising: a first plate, a second plate, a spacer, and an adapter, wherein: i. The two plates are movable relative to each other into different configurations; ii. One or both of the plates are flexible; iii. Each of the two plates includes an inner surface having a sample contact area for contacting a fluid sample; iv. One or both of the two plates include the spacer permanently fixed on the sample contact area of the corresponding plate; v. The spacer has: (a) a predetermined uniform height having a value selected in the range of 1.5 µm to 30 µm, (b) a column shape having a uniform cross-section and a flat top surface; (c) a ratio of the width of the spacer to the height equal to or greater than 1; (d) a predetermined fixed, non-random spacer spacing in the range of 10 µm to 200 µm; (e) a filling factor equal to 3% or greater, where the filling factor is the ratio of the contact area of the spacer fixed on the plate to the total area of the plate; and (f) the product of the filling factor and the Young's modulus of the spacer is 2 MPa or greater; vi. One or both of the two plates include a reagent coated on the sample contact area of the corresponding plate; vii. The reagent has at least one of the following: (a) a component that makes the PLT distribution uniform; (b) a component that reduces the aggregation of the PLT; (c) a component that stains the PLT; (d) a component that dilutes the blood; viii. The adapter includes: (a) a housing, (b) an attachment member on the housing that allows the adapter to be attached to a mobile phone having a camera, (c) a slot in the housing that allows (1) the plates in the closed configuration to slide into the slot, and (2) when the two plates are in the slot, at least a part of the sample area is less than 2 cm from the outer surface of the camera; wherein one of the configurations is an open configuration, in which the two plates are partially or completely separated, the spacing between the two plates is not adjusted by the spacer, and the sample is deposited on one or both of the plates; and wherein the other of the configurations is a closed configuration configured after the sample is deposited in the open configuration; and in the closed configuration, at least a part of the sample is compressed by the two plates into a layer with a very uniform thickness and is stationary relative to the two plates, where the uniform thickness of the layer is limited by the sample contact areas of the two plates and is adjusted by the two plates and the spacer.

159. The device according to claim 158, wherein the spacer height, the spacing between the two plates, and / or the sample thickness is 5 µm. ​ ​ 160. The device according to claim 158, wherein, the spacing between the two plates and / or the thickness of the sample is 0.8 µm to 10 µm.

161. The device according to claim 158, wherein, the spacer is a rounded rectangle.

162. The device according to claim 158, wherein, the lateral dimension of the spacer is 30 µm to 40 µm.

163. The device according to claim 158, wherein, the lateral dimension of the spacer is 5 µm to 50 µm.

164. The device according to claim 158, wherein, the rounded diameter of the spacer is 5 µm to 15 µm.

165. The device according to claim 158, wherein, the spacer is an array of rectangular dots.

166. The device according to claim 158, wherein, the spacing pitch of the spacer is 80 µm.

167. The device according to claim 158, wherein, the spacing pitch of the spacer is 70 µm to 150 µm.

168. The device according to claim 158, wherein, the length of one plate is 27 mm and the width of this plate is 22 mm.

169. The device according to claim 158, wherein, the length of one plate is 32 mm and the width of this plate is 24 mm.

170. The device according to claim 158, wherein, The area of one plate is 600 mm 2 and the area of another plate is 750 mm 2 .

171. The device according to claim 158, wherein, the thickness of one plate is 175 µm.

172. The device according to claim 158, wherein, the thickness of one plate is 1 mm.

173. The device according to claim 158, wherein, a reagent is dissolved in more than 70% alcohol for coating.

174. The device according to claim 173, wherein, the reagent coating uses the wetting property between the plate and the reagent solvent.

175. The device according to claim 173, wherein, the reagent is coated by spraying.

176. The device according to claim 158, wherein, Zwittergent is coated onto the first plate or the second plate or both.

177. The device according to claim 158, wherein, Acridine orange is coated on the two plates at an area concentration of 20 to 80 ng / mm 2 and Zwittergent is coated on the plates at an area concentration of 30 to 120 ng / mm 2 of area concentration.

178. The device according to claim 177, wherein, the material at the coating position of the first plate and the coating position of the second plate is polymethyl methacrylate.

179. The device according to claim 158, wherein, the landing mark for the blood droplet is on the outer surface of the first plate or the second plate.

180. The device according to claim 179, wherein, the landing mark for the blood droplet is a small dot or a small cross.

181. The device according to claim 179, wherein, the landing mark for the blood droplet is outside the field of view of the image.

182. The device according to claim 179, wherein, the landing mark for the blood droplet is near the center of the two plates.

183. The device according to claim 158, wherein, a separate paper or board between two boards for protecting the reagent coating on the board.

184. The device according to claim 158, wherein, a separate paper or board between two boards for picking up and opening the device.

185. The device according to claim 158, wherein, having a filling mark for notifying the user of the minimum volume and coverage area of the sample in the device.

186. The device according to claim 158, wherein, at least one of the two boards is transparent.

187. The device according to claim 158, wherein, the adapter has a band - pass excitation filter with a transmission band from 450 nm to 480 nm.

188. The device according to claim 158, wherein, the adapter has a long - pass emission filter with a cut - off wavelength between 510 nm and 525 nm.

189. The device according to claim 158, wherein, the adapter has an excitation light irradiation angle greater than the collection angle of the imaging optics.

190. The device according to claim 158, wherein, the adapter has a focal length between 2 mm and 4 mm.

191. The device according to any one of claims 1 - 3, 54, 95, 127, 158 for analyzing a blood sample, measuring the amount of a target analyte, or performing a WBC count.

192. The device according to any one of claims 1 - 3, 54, 95, 127, 158, wherein, the spacer height is less than 10 mm.

193. The device according to any one of claims 1 - 3, 54, 95, 127, 158, wherein, the spacer height is less than 200 µm.

194. The method according to any one of claims 4 - 6, wherein, depositing the sample in an open configuration onto one or two boards; then pressing the two boards into a closed configuration such that at least a portion of the sample is compressed into a layer with a very uniform thickness, the layer with a very uniform thickness being stationary on the board and defined by the inner surfaces of the two boards.

195. The device according to any one of claims 2 - 3, 54, 95, 127, 158, wherein, when in the closed configuration, a camera can be used to capture an image of the sample layer; the camera can have a field of view, which is defined as the area where the camera can capture an image of the sample; the camera is part of the device; the mobile phone is a mobile communication device, a tablet computer, or a laptop computer; the camera has one lens or two lenses aligned parallel to each other.

196. The device according to any one of claims 1 - 3, 54, 95, 127, 158, wherein, Different spacer heights can affect the accuracy of cell counting; the spacer height and the FoV affect the accuracy and consistency of the counting results; with an acceptable level of consistency, the direct counting results can be adjusted to reflect the true number of cells.

197. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The thickness of the layer of uniform thickness is in the range of 2.0 µm to 7.5 µm.

198. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The thickness of the layer of uniform thickness is in the range of 7.5 µm to 10.5 µm.

199. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The thickness of the layer of uniform thickness is in the range of 9.5 µm to 12.5 µm.

200. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The thickness of the layer of uniform thickness is in the range of 9.5 µm to 12.5 µm.

201. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The thickness of the layer of uniform thickness is in the range of 11.5 µm to 13.5 µm.

202. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The thickness of the layer of uniform thickness is in the range of 12.5 µm to 14.5 µm.

203. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The thickness of the layer of uniform thickness is in the range of 13.5 µm to 16 µm.

204. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The spacer height is in the range of 5.0 µm to 8.5 µm.

205. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The spacer height is in the range of 7.5 µm to 10.5 µm.

206. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The spacer height is in the range of 9.5 µm to 12.5 µm.

207. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The spacer height is in the range of 9.5 µm to 12.5 µm.

208. The device according to any one of claims 1 to 3, 54, 95, 127, 158, wherein, The spacer height is in the range of 11.5 µm to 13.5 µm.

209. The device according to any one of claims 1 to 3, 54, 95, 127, 158, Among them, the height of the spacer is in the range of 12.5 µm to 14.5 µm.

210. The device according to any one of claims 1 to 3, 54, 95, 127, 158, Among them, the height of the spacer is in the range of 13.5 µm to 16 µm.

211. The device according to any one of claims 1 to 3, 54, 95, 127, 158, Among them, The field of view for counting and differentiating WBCs is 0.1 mm 2 , 10 mm 2 , 50 mm 2 or 100 mm 2 .

212. The device according to claim 195, Among them, When the gap size between the plates is 10 µm, the FoV is greater than 36 mm 2 , whereby the WBC count and discrimination accuracy are less than 5%.

213. The device according to claim 195, Among them, When the gap size between the plates is 10 µm, the FoV is greater than 16 mm 2 , whereby the WBC count and discrimination accuracy are less than 10%.

214. The device according to claim 195, Among them, When the gap size between the plates is 10 µm, the FoV is greater than 2 mm 2 , whereby the WBC count and discrimination accuracy are less than 20%.

215. The device according to claim 195, Among them, When the gap size between the plates is 30 µm, the FoV is greater than 10 mm 2 , whereby the WBC count and discrimination accuracy are less than 15%.

216. The device according to claim 195, Among them, When the gap size between the plates is 30 µm, the FoV is greater than 50 mm 2 , whereby the WBC count and discrimination accuracy are less than 10%.

217. The device according to claim 195, Among them, The field of view is between 0.1 mm 2 and 10 mm 2 The gap size between the plates is in the range of 10 µm to 30 µm, or 30 µm to 50 µm, and the counting and discrimination accuracy is less than 10%.

218. The device according to claim 195, Among them, The field of view is between 0.1 mm 2 and 10 mm 2 in range, and the gap size between the plates is between 10 µm and 30 µm, whereby the counting and discrimination accuracy is less than 20%.

219. The device according to claim 195, Among them, The field of view is between 10 mm 2 and 50 mm 2 The gap size between the plates is in the range of 5 µm to 30 µm or 10 µm to 30 µm, and the counting and discrimination accuracy is less than 10%.

220. The device according to claim 195, Among them, The field of view is within 10 mm 2 to 50 mm 2 In the range, the gap size between the plates is in the range of 2 µm to 5 µm, 5 µm to 10 µm, or 10 µm to 30 µm, and thus the counting and discrimination accuracy is less than 20%.

221. The device according to claim 195, Among them, The field of view is within 50 mm 2 to 100 mm 2 and the gap size between the plates is within the range of 2 µm to 5 µm, 5 µm to 10 µm, 10 µm to 30 µm or 30 µm to 50 µm, whereby the counting and discrimination accuracy is less than 10%.

222. The device according to any one of claims 1 to 3, 54, 95, 127, 158 Among them, the height range of the spacer is 2 µm to 5 µm, whereby the WBC missing count is less than 15%.

223. The device according to claim 195, Among them, the height range of the spacer is 2 µm to 5 µm or 5 µm to 10 µm, whereby the WBC missing count is less than 30%.

224. The device according to claim 195, Among them, the height range of the spacer is 2 µm to 5 µm, 5 µm to 10 µm or 10 µm to 30 µm, whereby the WBC missing count is less than 60%.

225. The method according to claim 39, Among them, the reagent is coated in an array by droplet printing.

226. The method according to claim 39, Among them, the reagent is coated by spraying.

227. The method according to claim 39, Among them, the reagent is dissolved in more than 70% alcohol for coating.

228. The method according to claim 227, Among them, the coating uses the wetting property between the plate and the reagent solvent.

Images