System for examining biological samples under controlled environmental conditions

Abstract

Systems for examining biological samples under controlled environmental conditions, including apparatus and methods.

Description

[0001] Cross-references to related applications This application is based on and claims the benefits of U.S. Provisional Patent Application Serial No. 63 / 583580, filed September 18, 2023, pursuant to 35 USC § 119(e). This application incorporates, in its entirety, U.S. Provisional Patent Application Serial No. 63 / 583580, filed September 18, 2023; U.S. Patent No. 11,041,806, granted June 22, 2021; and U.S. Patent Application Publication No. US-2016-0123886-A1, published May 5, 2016. The systems described herein can be used in conjunction with the systems, features, and determinations described in the incorporated patent documents. Background Technology

[0002] Systems for analyzing biological samples are useful in medical research, life sciences, and other applications. Existing systems for measuring the properties of such samples (alone or in combination with factors of action such as candidate drug compounds) typically consist of an array of wells (such as microplates) with cells added to each well and a supporting material. The wells can then be analyzed serially or in parallel, depending on the assay design and the instrument used. Importantly, many of these assays are performed under non-physiological conditions, such as at room temperature rather than physiological temperatures, which can potentially affect the results. Therefore, there is a need for systems that can examine biological samples under controlled environmental conditions. Summary of the Invention

[0003] This disclosure provides a system for examining biological samples under controlled environmental conditions, including apparatus and methods. Attached Figure Description

[0004] Figure 1 This is a high-level schematic diagram of an exemplary system for examining biological samples under controlled environmental conditions, according to aspects of this disclosure. The schematic diagram discloses three alternative locations for an environmental control room (ECC): (A) external, (B) internal, in a non-inspection area, and (C) internal, in an inspection area.

[0005] Figure 2 is Figure 1 A pair of isometric views of an embodiment of the system, excluding portions of the environmental control room and housing, show (A) a view from above the front corner of the system and (B) a view from above the opposite rear corner of the system, highlighting the irradiation module, detection module, controller, and housing.

[0006] Figure 3 This is a top view of the system in Figure 2.

[0007] Figure 4 This is a side elevation view of the system in Figure 2.

[0008] Figure 5 is a schematic side view of a first exemplary system for examining biological samples under controlled environmental conditions, wherein the system has an external environmental control chamber, showing the configuration before (A), during (B), and after sample examination. This system corresponds to... Figure 1 Alternative option (A) is provided.

[0009] Figure 6 is a schematic side view of a second exemplary system for examining biological samples under controlled environmental conditions, wherein the system has an internal environmental control chamber located at a different location from the sample examination site, showing the configuration before (A), during (B), and after the examination. This system corresponds to... Figure 1 Alternative option (B) is provided.

[0010] Figure 7 is a schematic side view of a third exemplary system for examining biological samples under controlled environmental conditions, wherein the system has an internal environmental control chamber located at the sample examination site, showing the configuration before and during (A) the examination, and after (B) the examination. This system corresponds to... Figure 1 Alternative option (C) is provided.

[0011] Figure 8 This is a schematic side view of an environmental control room with integrated optical components.

[0012] Figure 9 This is a schematic side view of an environmental control room with an integrated glass heater.

[0013] Figure 10 This is a schematic side view of an environmental control room with integrated optics and glass heaters.

[0014] Figure 11 Figure 2 to Figure 4 A partial cross-sectional side elevation view of the system's detection module, including Figure 10 The environmental control room. Detailed Implementation

[0015] This disclosure provides a system for examining biological samples under controlled environmental conditions, comprising apparatus and methods. Control can be performed before, during, and / or after the examination. The system may include an irradiation module, a detection module, and an environmental control chamber. The irradiation module is configured to generate irradiation and direct irradiated light to the examination site. The detection module is configured to detect response light generated by one or more biological samples located at the examination site in response to irradiation by the irradiated light. The environmental control chamber is configured to control the environmental conditions of the biological samples before, during, and / or after the examination (in the last case, before further examination, etc.).

[0016] Figure 1 This is a high-level schematic diagram of an exemplary system 20 for examining a biological sample under environmental control, according to aspects of this disclosure. The system includes an irradiation module 22, a detection module 24, and an environmental control chamber (EEC) 26. A biological sample 28 (which may include one or more cells 29) can be positioned in a suitable sample holder 30. Some or all of the system may be located within a housing 31. The sample holder and the sample can be positioned at an examination point 32 for analysis. When the biological sample is at the examination point, irradiation light 34 generated by the irradiation module can be directed onto the biological sample, and response light 36 from the biological sample can be directed onto (or into) the detection module. The environmental control chamber is shown relative to the housing in three alternative mounting positions: (A) external, (B) internal, in a non-inspection area, and (C) internal, in an inspection area. These are respectively indicated by bidirectional horizontal arrows (…). ) a and b As shown, the sample holder and the biological sample it contains can be moved between the external (A) or internal (in the non-inspection area) (B) environmental control room and inspection point 32 via a stage or other mechanism. Controller 38 can operate or otherwise control one or more components of the system.

[0017] An illumination module can be used to generate illumination light capable of monitoring status and / or eliciting a response from the sample. This module may include one or more individual light sources (e.g., one, two, three, four, five, six, seven, eight, nine, or more sources). Light sources may include lasers, light tubes, and / or light-emitting diodes (LEDs), etc. Lasers may include single-mode lasers and / or multimode lasers. Each light source may be capable of emitting at one or more dominant single wavelengths (e.g., 488 nm or 514 nm) or within one or more wavelength ranges (e.g., 450 nm to 550 nm). In some cases, two or more light sources may output light with the same spectral properties, where light from two or more sources is combined to increase its intensity. In other cases, two or more light sources may output light with different spectral properties to broaden the range of available illumination wavelengths, allowing the light engine to be used with a wider range of samples and a greater number of samples. The number of light sources (or groups of light sources) may be independent of or proportional to the number of samples and / or sample sites being examined. For example, each sample or sample spot may have one light source (or a group of light sources): one light source (or a group of light sources) for every two samples or sample spots, one light source (or a group of light sources) for every four samples or sample spots, and so on. The intensity of the light from each light source can be adjusted independently, for example, from 0% to 100% relative intensity. During a specific analysis or determination, the light output by the irradiation module may come from a single source or may be a mixture of light from two or more sources. The spectral characteristics of the light output by the light engine can be matched to its intended use, for example, to activate photoexciteable compounds and / or processes, and / or to excite fluorescence from preselected fluorescent labels, etc. Depending on the requirements of the specific activation and / or inspection protocol, multiple wavelengths of light may be applied simultaneously or sequentially. The irradiation module may optionally include diffusers, speckle removers, and / or other mechanisms for reducing inhomogeneities in the irradiation light. The irradiation module may include reflective and / or refractive elements for combining light from different light sources into a single optical path. The irradiation light can irradiate all or part of a biological sample during use. When a sample is contained in a multi-well plate, illumination light can simultaneously illuminate one, some, many, or all of the wells of the plate. Illumination light is particularly suitable for photostimulation of cellular responses and recording of cellular responses. The illumination module may include one or more lenses or lens arrays and directs the illumination light to one or more lenses or lens arrays that collect and focus the light onto one or more biological samples.

[0018] The detection module can be used to capture the response light generated by a biological sample in response to illumination by an irradiation light, and to produce a corresponding signal (or multiple signals) or other manifestations of the response light for further analysis. The biological sample and / or array of biological samples may have a non-zero spatial range transverse to the irradiation direction, and / or include a series of discrete samples that again have a non-zero spatial range. Therefore, the detection module may be able to detect the response light at a range of transverse positions. The detection module may include a multi-element array of point detectors, such as photodiodes or photomultiplier tubes (PMTs). The point detectors may be spatially registered 1:1 to the orifices of a porous sample holder, wherein one or more lenses or lens arrays are used to focus the response light from each orifice onto its corresponding detector. Alternatively or additionally, the detection module may include an imaging detector, such as a camera, including complementary metal-oxide-semiconductor (CMOS) cameras, scientific CMOS (sCMOS) cameras, and / or charge-coupled device (CCD) cameras, etc. Signals from multiple points in the sample or from multiple samples may be collected simultaneously by all elements of the array detector or all portions of the imaging detector (i.e., through parallel sampling). Signal acquisition can be fast (e.g., from about 10 Hz to about 10 kHz). High sampling rates provide the temporal resolution needed to accurately track cellular responses on a millisecond timescale.

[0019] An environmental control chamber can be used to control one or more environmental conditions of a biological sample, such as temperature, gas composition, humidity, and / or pressure. These conditions can be kept substantially constant. For example, the temperature can be maintained at or near physiological temperature, such as 37°C. Alternatively or additionally, conditions can be varied (e.g., gradually increased, cyclically changed, or otherwise altered between different values). For example, the temperature can be increased or decreased from a baseline value to assess the effects of heating or cooling on the sample. The environmental control chamber may include ports (such as a lid and associated openings) for introducing and removing a sample holder and associated biological sample into and from the chamber. The environmental control chamber may further include sensors for monitoring one or more environmental conditions within the chamber, and actuators for bringing about desired changes to the monitored conditions. For example, the chamber may include a temperature sensor for measuring temperature, and one or more heaters or coolers for raising or lowering the temperature within the chamber, respectively. The actuators may be integrally integrated into the chamber, such as heating or cooling plates, or may be external, such as an infrared light source in an irradiation module that generates infrared light for absorption by the sample and / or associated sample holder. In the latter case, a filter can be used between the sample and the detector to reduce or eliminate infrared radiation before it becomes detectable and interferes with the response light. The chamber may further include or be associated with optics for focusing or otherwise directing the illumination light onto the biological sample and for collecting and directing the response light generated by the sample to the detection module.

[0020] Biological samples may include any particles, substances, extracts, mixtures, and / or assemblies derived from or corresponding to one or more organisms, cells, and / or viruses. Biological samples may also include additional components to facilitate analysis, such as fluids (e.g., water), buffers, culture nutrients, salts, other reagents, dyes, etc. Biological samples will typically include cells. These cells may be set in any suitable medium, such as culture medium. In some cases, the cells may be adherent cells, which preferentially or completely attach to a substrate (such as glass or plastic) and / or be in contact with the substrate for culture. Suitable cells may include cells derived from animals, plants, bacteria, and / or fungi (including yeast), etc. Cells may be primary cells, immortalized cells, senescent cells, stem cells, differentiated cells, transformed cells, infected cells, transgenic cells, cells assembled into tissues or multicellular organisms (such as embryos), and / or similar cells. Substances as used herein may include any small or large compounds isolated from (or secreted by) cells. Exemplary substances may include proteins (e.g., enzymes, receptors, binding partners, ion channels, etc.), peptides, nucleic acids (such as cyclic or acyclic mononucleotides, DNA and / or RNA), carbohydrates, lipids, ions, hormones, metabolites, their conjugates, and / or analogs. Extracts and / or assemblies as used herein may include fractions or organelles isolated from cells. Exemplary fractions and / or organelles may include crude or at least partially purified cell nuclei, cell membranes, mitochondria, cytosols, whole-cell lysates, etc. Assemblies may also include 3D spheres. Samples may be selected to facilitate screening for the biological activity (including the absence of (adverse) activity) of compounds (such as drug candidates) during safety screening. For example, samples may be selected to facilitate rapid screening of pharmacologically active compounds targeting ion channels or for safety pharmacological assays. Exemplary cells include cardiomyocytes, etc. For simplicity, the term "sample" as used herein may refer to a single sample or multiple samples depending on the context. For example, the phrase "sample holder and sample" refers to a single sample if the sample holder is configured to hold a single sample, or to multiple samples if the sample holder (such as a multi-well plate) is configured to hold (and carry) multiple samples.

[0021] Biological samples may be supported or otherwise held by a suitable sample holder. A sample holder generally comprises any substrate or other mechanism for holding the sample for irradiation and detection. A sample holder may hold one or more discrete samples at one or more different sample sites. In some cases, the sample sites may be defined by a mechanical barrier (e.g., a wall) to form sample wells. In other cases, sample sites may be defined by a chemical barrier, such as a hydrophobic region separating a hydrophilic region, or spatially separated binding sites for nucleic acids, proteins, and / or other materials. Sample sites may be individual fluid volumes or share a common fluid volume. Exemplary sample holders with individual fluid volumes may include multiwell plates, such as microplates and PCR plates. Such plates may have any suitable number of sample wells, such as 4, 6, 12, 24, 48, 96, 384, or 1536 sample wells. Exemplary sample holders with a common fluid volume may include microscope slides and / or sample chips for nucleic acids and / or proteins, etc. Depending on the analysis, samples can in all cases be independent of each other, or equal fractions or replicates. Samples may include control samples or calibration samples. The sample holder may be at least partially transparent to both the illumination light and the examination light, at least in the main direction of light propagation during the examination.

[0022] The stage may include any structure configured to support and move a sample holder and associated biological sample for input, output, and / or inspection. It may allow samples to be added to and removed from the system (e.g., at an input / output area). It may also transport samples between the environmental control room and the inspection point (when these are not contiguous). The stage may include rails, belts, gears, pulleys, and motors, etc., with its movement controlled by a controller.

[0023] The controller may include any mechanism for assessing the status of the system (e.g., the location, identification, and / or status of samples), for operating optics (e.g., determining and executing irradiation schemes and / or collecting the generated response light), and for transporting sample holders and samples between input / output areas, environmental control rooms, and inspection points (if different). The controller may also monitor and report the status of the system itself and its sub-components.

[0024] The system may further include various optical elements or devices for performing corresponding purposes. These optical devices may be directly associated with one or more main system components, such as an illumination module, a detection module, and an environmental control chamber. These optical devices may also be distributed among system components as needed. Examples of optical devices include mirrors, lenses, beam splitters, spectral filters (such as fluorescence excitation and emission filters), and neutral density filters. Optical devices can act on illumination light, response light, and / or any additional light (such as infrared radiation used to heat the sample). Mirrors use reflection to change the direction and / or focus of light, allowing, among other uses, the combination of light from different sources and folding the optical path through the system to reduce its footprint. Lenses use refraction to bend light, allowing light to be focused, collimated, or diverged. This is particularly important when illumination light is directed onto the sample and response light is collected from the sample and directed onto the detector. Beam splitters separate light, typically causing some light to transmit along one path and the remainder to reflect along another. Separation may depend on the spectrum of light, or it may not (e.g., in the former case, shorter wavelengths are reflected and longer wavelengths are transmitted). Spectral filters can partially or completely block light with some wavelengths and partially or completely transmit light with other wavelengths. These filters play an important role in “cleaning up” the light from the irradiation module (e.g., by reducing or eliminating light with undesired wavelengths before it reaches the sample). They can also play an important role in fluorescence measurements. Filters placed before the sample (called excitation filters) reduce or remove wavelengths of light unwanted for exciting fluorescence (especially if these wavelengths can be confused with fluorescence emission light). Filters placed after the sample (called emission filters) reduce or remove wavelengths of light not associated with fluorescence emission (especially wavelengths corresponding to stray excitation light) to help ensure that the detected light is the responsive light by reducing contamination. Emission filters are particularly important when the excitation light and the collected emission light follow the same optical path (i.e., in a transmission fluorescence versus epifluorescence configuration). A neutral density filter reduces the intensity of light (including illumination and / or response light) substantially uniformly (neutrally) across at least the wavelength of interest. This can improve performance if the intensity were originally too high. Alternatively or additionally, the intensity of the illumination light (and therefore indirectly, the response light) can be controlled by the illumination module itself, for example, by varying the intensity and / or duration of the power supplied to the light source. The relative options, positions, sizes, and / or other properties of these various optics can be selected to increase or decrease the portion of the sample holder (and therefore the sample) being illuminated, and / or to alter the quality of the illumination.

[0025] Lenses can play a particularly important role in the system and can be positioned within the path taken by the illumination light, the path taken by the response light, or both paths. These lenses can perform any suitable function. For example, a lens positioned in the illumination path can homogenize and collimate the illumination light incident on the sample holder, making its intensity more uniform and / or closer to being parallel to the optical axis and / or perpendicular to the plane of the sample holder to reduce shading. Alternatively or additionally, a lens positioned in the response path can collect the response light and direct it toward the detection module to increase the amount of light captured by the detection module. Lenses can also focus light onto the detection module to ensure that the position of the light on the detector is better correlated with its origin position in the sample. Lenses positioned between the main components of the system can complement or enhance the function of lenses integrated with the illumination module, detection module, and / or environmental control chamber. Depending on the position and function of the lens, it may be referred to differently and not limited to condenser lenses, objectives, field lenses, and / or tube lenses. These lenses can also operate in pairs or in triplets (etc.) along a given path, and / or as lens arrays (etc.) operating laterally to the illumination and response path.

[0026] Figure 2 to Figure 4 It shows Figure 1 Various views of a portion of embodiment 50 of the system. This portion includes an irradiation module 52, a detection module 54, a controller 56, and a housing 58. Figure 1 The three possible locations A, B, and C of the environmental control room are shown and described in Figures 5 through 7, and are also described in the relevant text.

[0027] Figure 5 is a schematic side view of a first exemplary system 70 for examining biological samples. Here, an environmental control chamber 72 is located outside a housing 74, which houses other components of the system used for examination. Figure 5 illustrates three exemplary configurations associated with sample examination. (A) Prior to examination, a sample holder 76 containing the biological sample is positioned within the environmental control chamber and held there until the examination is performed. Temperature and / or other environmental conditions are applied and / or changed. The sample is then moved to the examination location by opening a door 78 separating the environmental control chamber and the housing, and using a stage 80 to transport the sample holder and sample through the door to the examination point. (B) During examination, the door between the environmental control chamber and the housing is closed, the sample holder is positioned at the examination point, the sample is illuminated with light, and the resulting response light is detected. (C) After examination, the door between the housing and the environmental control chamber is reopened, and the sample holder and sample are transported back to the environmental control chamber. The same sample can be analyzed again by repeating the steps associated with configurations A and B. Alternatively, as shown, the cover 82 or other closing mechanism of the environmental control chamber can be opened, and the sample holder and sample can be removed. The entire process can be repeated for a new sample. Depending on the embodiment, the environmental control chamber here and elsewhere in this disclosure may house a single sample holder or multiple sample holders. In the latter case, the stage will select the sample holder and sample of interest based on instructions from the controller.

[0028] Figure 6 is a schematic side view of a second exemplary system 90 for examining biological samples. Here, an environmental control chamber 92 is located inside a housing 94, which houses other components of the system used for examination. However, the location of the environmental control chamber differs from and is distinct from the examination point 96 (dashed rectangle) for examining the sample. Figure 6 illustrates three exemplary configurations associated with sample examination. (A) Before examination, a sample holder 98 containing the biological sample is positioned within the environmental control chamber. Temperature and / or other environmental conditions are applied and / or changed. The sample is then moved to the examination point for examination by moving the sample holder using a stage 100. (B) During examination, the sample is irradiated with light, and the resulting response light is detected. (C) After examination, a door 102 in the housing is opened, and the sample holder is transported to the outside by the stage and removed. The same sample can be analyzed more than once by repeating the steps associated with configurations A and B before performing the steps associated with configuration C. Alternatively, once the steps associated with construction C have been completed, a new sample holder can be placed on the stage, the sample holder can be transported to the environmental control room, the door in the housing can be closed, and the new sample can then be examined as described above. Removing the sample from the environmental control room during examination reduces the complexity of the control room.

[0029] Figure 7 is a schematic side view of a third exemplary system 110 for examining biological samples. Here, similar to Figure 6, an environmental control chamber 112 is located inside a housing 94 that houses other components of the system for performing the examination. However, in this embodiment, the examination point 116 (dashed rectangle) is located inside the environmental control chamber, not outside. Therefore, the sample can be examined while it is inside the environmental control chamber to ensure better control of sample conditions, while increasing the complexity of the environmental control chamber (which must now be at least partially transparent). Figure 7 illustrates two exemplary configurations associated with sample examination. (A) Before and during examination, a sample holder 118 is positioned at the examination point in the environmental control chamber. Temperature and / or other environmental conditions are applied and / or changed. The sample is irradiated with light, and the resulting response light is detected. The sample can be examined at a single time or time interval, or at different times or time intervals, all without leaving the environmental control chamber. (B) After examination, a door 120 in the housing is opened, and the sample holder is transported to the outside and removed using a stage 122. The new sample holder can then be placed on the stage, transported to the environmental control chamber, the door in the housing closed, and the new sample can be inspected as described above. A suitable environmental control chamber for this embodiment is... Figures 8 to 10 As shown in the image.

[0030] Exemplary details of suitable irradiation modules, detection modules, controllers, and housings for the embodiments shown in Figures 5 to 7 are shown in Figures 2 to 7. Figure 4 As shown in the present disclosure and in other locations, and further described in cross-referenced patent documents.

[0031] Figure 8 This is a schematic side view of an environmental control chamber 130 with integrally integrated optics. The control chamber includes a central area 132 having a controlled environment and sized to receive at least one sample holder. The central area is disposed between two walls that are at least substantially planar and at least substantially parallel. A first illumination wall 134 is configured to transmit illumination light 135 from an illumination module toward the sample holder and associated biological sample housed within the chamber. A second detection wall 136 is configured to transmit response light 137 generated by the biological sample to the detection module. Here, the illumination and detection walls can be formed using any material capable of transmitting all or all of the illumination and detection light. Suitable materials may include transparent plastics and / or glass. The walls may then include suitable illumination optics 138a and detection optics 138b, such as lens arrays. The number of lenses or microlenses in the lens array may be matched to the number of samples or sample holes in the sample holder, such that each lens illuminates (138a) a single sample or hole and / or detects (138b) from a single sample or hole. The optics may be molded into plastic.

[0032] Figure 9 This is a schematic side view of an environmental control room 150 with an integrated glass heater. Environmental control room 150 and... Figure 8 The environmental control chamber 130 shown shares many similarities. For example, similar to control chamber 130, control chamber 150 includes a central area 152 with a controlled environment and sized to receive at least one sample holder. Furthermore, the central area is positioned between two walls that are at least substantially flat and at least substantially parallel: an illumination wall 154 configured to transmit illumination light 155 and a detection wall 156 configured to transmit detection light 157. However, in this embodiment, one or both walls include optically transparent heating elements 158a, b. The walls may include transparent glass, and the heating elements may include an indium tin oxide (and / or other electrically heatable) coating. Resistance heating occurs if the coating is deposited in a meandering path (e.g., a serpentine path and / or a sawtooth path, etc.) and a voltage difference is applied between one end of the path and the other. This heating can serve as a heat input to the environmental chamber to allow for the establishment and / or maintenance of temperature. Furthermore, since indium tin oxide reflects infrared light, it can be used as both a filter and a heater.

[0033] Figure 10 It is an optical device with integrated components (such as...) Figure 8 ) and glass heaters (such as Figure 9A schematic side view of an environmental control chamber 170. The environmental control chamber 170 further includes a central area 172 having a controllable environment, sized to receive at least one sample holder; and parallel irradiation walls 174 and 176, as defined above, which are at least substantially planar. Furthermore, as described above, one or both walls include both integrally integrated optics and integrally integrated heating elements. The integrally integrated features on the upstream (irradiation or excitation) side of the environmental control chamber may include one or more of an irradiation light collimating optics 174a, an irradiation light filter 174b, an irradiation light focusing lens element array 174c, and an irradiation light-side heating element. These integrally integrated features collimate the incident irradiation light 175 to 174a, filter 174b light to remove unintended or undesirable wavelengths and / or reduce intensity, focus light 174c onto the sample, and control the temperature 174d within the sample chamber. The integrated features on the downstream (detection or emission) side of the environmental control chamber may include one or more of the following: detection light collimating optics 176a (such as an array of emission light collimating lens elements), detection light filter 176b, detection light focusing lens element array 176c, and detection-side heating element 176d. These integrated features collimate the response light 177 to 174a, filter the response light 174b, and then refocus the filtered light 174c onto the sensor or sensor array in the detection module, while again controlling the temperature 176d of the sample chamber. The aspects of the environmental control chamber (including the various optics and heating elements) can be arranged in any suitable order, but the heating elements are typically positioned closest to the sample holder.

[0034] Figure 11 Figure 2 to Figure 4 The detection module (or read head) 54 (including Figure 10A partial cross-sectional side elevation view of the environmental control room 170. The optical path is generally indicated by arrows, showing the excitation (irradiation) light 185 and the emission (detection) light 187. This figure shows the detection module in use, where the porous sample holder 190 and the associated sample orifice / sample 191 are configured for inspection within the environmental control room 192. Upper heating elements 194a and lower heating elements 194b are positioned above and below the sample holder on the upstream excitation (irradiation) side and downstream emission (inspection) side of the inspection point and sample holder, respectively. The heating elements can be used to maintain and / or change the temperature of the sample holder and the associated sample. The excitation light 185 from the irradiation module is guided here toward the inspection point via a pair of 100% mirrors 196a, 196b. In this embodiment, the excitation light passes through an excitation collimating Fresnel lens 198 (which collimates the light), an excitation focusing lens array 200 (which focuses the collimated light toward discrete points (e.g., sample apertures) in the sample holder), an excitation filter 202 (which cleans the excitation light), and an upper heating element. Emitted (e.g., fluorescence) light 187 from the sample passes through a lower heating element, an emission objective array 204 (which collects and optionally collimates the emitted light), an emission filter 206 (which cleans the emitted light (particularly removing contaminants from the excitation light)), and an emission tube lens array 208 (which focuses the light onto a detector (e.g., detecting a printed circuit board assembly (PCBA) 210)). The detector detects the light and converts it into an electronic signal for processing by a controller. A stage 212 shuttles the sample holder and the associated sample back and forth to the inspection point.

[0035] Selected aspects This paragraph describes additional selected aspects of this disclosure, which are given in the form of a series of paragraphs without limitation, some or all of which may be numbered for clarity and efficiency. Each of these paragraphs may be combined with one or more other paragraphs in this application, and / or with disclosures elsewhere in this application, in any suitable manner. Some of the following paragraphs explicitly refer to and further limit the other paragraphs, providing examples of suitable combinations without limitation.

[0036] 1. A system for examining a biological sample, comprising: (A) an irradiation module configured to generate irradiation light and direct the irradiation light to an examination site; (B) a detection module configured to detect response light generated by the biological sample located at the examination site in response to irradiation by the irradiation light; and (C) an environmental control chamber configured to control the environmental conditions of the biological sample before and / or during examination of the biological sample.

[0037] 2. The system of paragraph 1 further includes a housing defining an inner side and an outer side, wherein the irradiation module, the detection module, and the inspection point are at least substantially disposed on the inner side of the housing.

[0038] 3. The system in paragraph 2, wherein the environmental control room is located outside the enclosure.

[0039] 4. The system in paragraph 3 further includes a platform for transporting biological samples between the environmental control room and the inspection point.

[0040] 5. The system in paragraph 2, wherein the environmental control room is located inside the enclosure.

[0041] 6. The system in paragraph 5, wherein the environmental control room is located outside the inspection point, further includes a platform for transporting biological samples between the environmental control room and the inspection point.

[0042] 7. The system in paragraph 5, in which the environmental control room is located at the inspection point.

[0043] 8. The system of paragraph 7, wherein the environmental control room has an at least partially transparent irradiation side through which irradiation light enters the environmental control room from the irradiation module; and an at least partially transparent detection side through which response light exits the environmental control room toward the detection module, wherein the irradiation side and the detection side are at least substantially planar and parallel to each other.

[0044] 9. The system of paragraph 8, wherein at least one of the irradiation side and the detection side includes a heating element.

[0045] 10. The system of paragraph 9, wherein the heating element is at least partially transparent and is positioned in the path of the irradiating light, the responding light, or both.

[0046] 11. The system of paragraph 10, wherein the heating element comprises an indium tin oxide coating.

[0047] 12. The system of any of paragraphs 7 to 11, wherein the environmental control room includes at least one lens disposed on one or both of the irradiation side and the detection side.

[0048] 13. The system of paragraph 12, wherein a biological sample is contained in a multiwell plate, and wherein an environmental control chamber includes a lens for each well in the multiwell plate.

[0049] 14. The system of paragraph 13, wherein the environmental control chamber includes at least two lenses for each well in the multi-well plate, one lens on the illumination side with respect to each well and one lens on the detection side with respect to each well.

[0050] 15. The system of paragraph 14, wherein the environmental control room further includes at least three lenses for each hole in the perforated plate, two lenses on one side of the perforated plate with respect to each hole, and at least one lens on the other side of the perforated plate with respect to each hole.

[0051] 16. The system of paragraph 14, wherein the environmental control chamber includes at least four lenses for each well in the multi-well plate, two lenses on the illumination side with respect to each well and two lenses on the detection side with respect to each well.

[0052] 17. A system of any of paragraphs 13 to 16, wherein the environmental control room includes at least one Fresnel lens on the irradiation side.

[0053] 18. A system of any of paragraphs 13 to 17, wherein the detection module collects response light from each aperture such that the intensity of light from each aperture is distinguishable from the intensity of light from each of the other apertures.

[0054] 19. A system of any of paragraphs 13 to 18, wherein the number of holes in the perforated plate is selected from the set of 4, 6, 12, 24, 48, 96, 384 and 1536.

[0055] 20. A system of any of paragraphs 12 to 19, wherein at least one lens is at least one Fresnel lens.

[0056] 21. Any system in the preceding paragraphs, wherein the environmental conditions are selected from a set of temperature and atmospheric composition.

[0057] 22. The system in paragraph 21, where the environmental condition is temperature.

[0058] 23. The system in paragraph 21, where the environmental condition is the concentration of carbon dioxide.

[0059] 24. Any system described in the preceding paragraph, wherein the illumination light is fluorescent excitation light and wherein the response light is fluorescent emission light.

[0060] 25. The system in paragraph 24, wherein the detection module determines the intensity of the fluorescent emitted light.

[0061] 26. The system in paragraph 25, wherein the biological sample consists of at least two discrete samples, wherein the detection module determines the intensity of fluorescence emission light with respect to each discrete sample.

[0062] 27. The system of any of paragraphs 24 to 26 further includes an excitation filter operatively disposed between the irradiation module and the inspection point, and configured to alter the spectral characteristics of the irradiation light before it is incident on the biological sample.

[0063] 28. The system of any of paragraphs 24 to 27 further includes an emission filter operatively disposed between the inspection point and the detection module, and configured to alter the spectral characteristics of the response light before the response light is incident on the detector.

[0064] 29. The system of paragraph 27 or 28, wherein the environmental control room is located at the inspection point, and wherein at least one of the excitation filter and the emission filter is integrally integrated with the environmental control room.

[0065] 30. Any system described in the preceding paragraphs, wherein the biological sample includes cells.

[0066] 31. The system in paragraph 30, in which the cells are cardiomyocytes.

[0067] 32. Any system described in the foregoing paragraphs further includes a controller that operates the irradiation module, the detection module, and, if present, a stage.

[0068] 33. The system in paragraph 32, wherein the controller further operates the environmental control room.

[0069] 34. The system in paragraph 32 or 33, wherein the controller instructs the detection module to detect the response light at a series of times as part of a kinetic determination.

[0070] 35. The system in paragraph 32 or 33, wherein the controller instructs the detection module to detect the response light in discrete time as part of the endpoint determination.

[0071] 36. Any system described in the preceding paragraphs further includes biological samples.

[0072] 37. The system in paragraph 36, in which biological samples include cells.

[0073] 38. The system in paragraph 37, in which the cells are cardiomyocytes.

[0074] 39. Any system described in the preceding paragraph, wherein the illumination light is fluorescent excitation light and the response light is fluorescent emission light, further comprising a fluorophore that reports the state of the biological sample.

[0075] 40. The system in paragraph 39, in which fluorophores report the activity of ion channels in biological samples.

[0076] 41. The system in paragraph 40, in which the ion is calcium.

[0077] 42. Any system described in the preceding paragraph, wherein the irradiation module is further configured to generate heating light for maintaining or increasing the temperature of the biological sample when the biological sample is positioned at the examination site, and wherein the intensity of the heating light and the intensity of the irradiation light can be changed independently.

[0078] 43. The system in paragraph 42, in which the heating light is infrared radiation.

[0079] 44. Any system described in the foregoing paragraphs further includes a temperature sensing element located at the inspection point.

[0080] 45. Any system described in the preceding paragraph, wherein the illumination module includes at least one LED, solid-state light source, or laser.

[0081] 46. ​​Any system described in the preceding paragraph, wherein the illumination module comprises multiple light sources, and wherein the intensity of light produced by each light source is independently controllable to allow for alteration of the spectral composition of the illumination light.

[0082] 47. Any system described in the preceding paragraph, wherein the detection module comprises an array of point detectors.

[0083] 48. Any system described in the preceding paragraph, wherein the detection module includes a camera.

[0084] 49. Any system described in the preceding paragraphs, wherein the sampling rate of the detection module is greater than approximately 100 Hz.

[0085] 50. A method for examining a biological sample, comprising: (A) selecting the biological sample; (B) selecting a system according to any of the preceding paragraphs; (C) irradiating the biological sample with illumination light; (D) detecting response light transmitted from the biological sample in response to the illumination light; and (E) determining the state of the biological sample using the response light.

[0086] 51. An environmental control room comprising: (A) an enclosed volume having at least a pair of substantially opposing walls, each of the opposing walls being at least partially transparent; and (B) a control mechanism configured to allow maintenance or alteration of environmental conditions within the room.

[0087] 52. The environmental control room in paragraph 51, in which the spectrum of light that can be transmitted through one of two opposing walls is different from the spectrum of light that can be transmitted through the other opposing wall.

[0088] 53. The environmental control room of paragraph 51 or 52, wherein at least one of the opposing walls further includes at least one lens for changing the focal point of light passing through the wall.

[0089] 54. The environmental control room of any of paragraphs 51 to 53, wherein at least one lens is a converging lens.

[0090] 55. An environmental control room of any of paragraphs 51 to 54, wherein at least one lens is a Fresnel lens.

[0091] 56. An environmental control room of any of paragraphs 53 and 54, wherein at least one lens is a regular array of lenses, each lens focusing light to a lateral position of the regular array in the sample plane and / or collecting light from a lateral position of the regular array in the sample plane.

[0092] 57. An environmental control room of any of paragraphs 51 to 56, wherein the control mechanism includes a at least partially transparent coating disposed on at least one of the opposing walls, the coating heating up when an electric current passes through it.

[0093] 58. The environmental control room in paragraph 57, wherein the coating includes indium tin oxide.

[0094] 59. The environmental control room of paragraph 57 or 58, wherein the control mechanism further includes electrical contacts for guiding electricity through the conductive coating.

[0095] 60. An environmental control room of any of paragraphs 51 to 59, wherein the control mechanism includes at least one port configured to receive a gas input.

[0096] 61. The environmental control room in paragraph 60 contains carbon dioxide.

[0097] 62. An environmental control room of any of paragraphs 51 to 61, wherein at least one of the opposing walls includes a spectral filter configured to alter the spectrum of light passing through the filter.

[0098] 63. The environmental control room of paragraph 62, wherein each of the opposing walls includes a spectral filter, and wherein the spectrum of light passing through each spectral filter is different from the spectrum of light passing through the other spectral filter.

[0099] As used in this disclosure, the term "and / or" means all combinations of the listed elements. For example, a list with two elements, "A and / or B," covers three possibilities: A only, B only, and both (A and B). Similarly, a list with three elements, "A, B, and / or C," covers seven possibilities: A only, B only, C only, both A and B, both A and C, both B and C, and all three (A, B, and C). The same pattern applies to extensions to four or more elements.

[0100] As used in this disclosure, the term “exemplary” means “illustrator” or “used as an example” and is not intended to imply preference or superiority.

[0101] The use of terms such as “first,” “second,” “A,” “B,” etc. in this disclosure is for the purpose of distinguishing or identifying the individual members in a set, and is based on the order introduced in the specific context and is not intended to indicate any order or quantity limitation.

[0102] The above disclosure may encompass multiple different inventions with independent uses. Although each of these inventions has been disclosed in its preferred form, the specific embodiments disclosed and illustrated herein should not be considered limiting, as many variations are possible. The subject matter of this invention includes all novel and non-obvious combinations and sub-combinations of the various elements, features, functions, and / or properties disclosed herein. The following claims specifically point to certain combinations and sub-combinations considered novel and non-obvious. Inventions embodied in other combinations and sub-combinations of features, functions, elements, and / or properties may be claimed in applications claiming priority to this application or related applications. Such claims, whether for different inventions or for the same invention, and regardless of whether their scope is broader, narrower, equal to, or different from the original claims, are also considered to be included within the subject matter of the invention disclosed herein.

Claims

1. A system for examining biological samples, comprising: An illumination module configured to generate illumination light and direct the illumination light to the inspection point; A detection module configured to detect response light generated by a biological sample located at the inspection point in response to irradiation by the irradiation light; as well as An environmental control room configured to control the environmental conditions of the biological samples before and / or during examination.

2. The system of claim 1, further comprising a housing defining an inner side and an outer side, wherein the irradiation module, the detection module, and the inspection point are disposed at least substantially within the housing.

3. The system according to claim 2, wherein, The environmental control room is located on the outside of the housing.

4. The system of claim 3, further comprising a stage for transporting biological samples between the environmental control room and the inspection point.

5. The system according to claim 2, wherein, The environmental control room is located inside the housing.

6. The system according to claim 5, wherein, The environmental control room is located outside the inspection point and further includes a platform for transporting biological samples between the environmental control room and the inspection point.

7. The system according to claim 5, wherein, The environmental control room is located at the inspection point.

8. The system according to claim 7, wherein, The environmental control room has an at least partially transparent irradiation side through which irradiation light enters the environmental control room from the irradiation module; and an at least partially transparent detection side through which response light exits the environmental control room toward the detection module, wherein the irradiation side and the detection side are at least substantially planar and parallel to each other.

9. The system according to claim 8, wherein, At least one of the irradiation side and the detection side includes a heating element.

10. The system according to claim 9, wherein, The heating element is at least partially transparent and is disposed in the path of the irradiation light, the response light, or both.

11. The system according to claim 10, wherein, The heating element includes an indium tin oxide coating.

12. The system according to claim 7, wherein, The environmental control room includes at least one lens, which is disposed on one or both of the irradiation side and the detection side.

13. The system according to claim 12, wherein, The biological sample is contained in a multi-well plate, and the environmental control chamber therein includes a lens for each well in the multi-well plate.

14. The system according to claim 13, wherein, The environmental control chamber includes at least two lenses for each hole in the porous plate, one lens on the illumination side with respect to each hole and one lens on the detection side with respect to each hole.

15. The system according to claim 14, wherein, The environmental control room further includes at least three lenses for each hole in the perforated plate, two lenses on one side of the perforated plate with respect to each hole, and at least one lens on the other side of the perforated plate with respect to each hole.

16. The system according to claim 14, wherein, The environmental control chamber includes at least four lenses for each hole in the porous plate, two lenses on the illumination side with respect to each hole, and two lenses on the detection side with respect to each hole.

17. The system according to claim 13, wherein, The environmental control room includes at least one Fresnel lens on the irradiation side.

18. The system according to claim 13, wherein, The detection module collects response light from each aperture, such that the intensity of light from each aperture can be distinguished from the intensity of light from each of the other apertures.

19. The system according to claim 13, wherein, The number of holes in the perforated plate is selected from the set of 4, 6, 12, 24, 48, 96, 384 and 1536.

20. The system according to claim 12, wherein, The at least one lens is at least one Fresnel lens.

21. The system according to claim 1, wherein, The environmental conditions are selected from a set of temperature and atmospheric composition.

22. The system according to claim 21, wherein, The environmental condition referred to is temperature.

23. The system according to claim 21, wherein, The environmental condition referred to is the carbon dioxide concentration.

24. The system according to claim 1, wherein, The illumination light is fluorescent excitation light, and the response light is fluorescent emission light.

25. The system according to claim 24, wherein, The detection module determines the intensity of the fluorescent emitted light.

26. The system of claim 25, wherein the biological sample comprises at least two discrete samples, wherein the detection module determines the intensity of the fluorescence emission light for each discrete sample.

27. The system of claim 24, further comprising an excitation filter operatively disposed between the irradiation module and the inspection point, and configured to alter the spectral characteristics of the irradiation light before the irradiation light is incident on the biological sample.

28. The system of claim 24, further comprising an emission filter operatively disposed between the inspection point and the detection module, and configured to alter the spectral characteristics of the response light before the response light is incident on the detector.

29. The system according to claim 27, wherein, The environmental control room is located at the inspection point, and at least one of the excitation filter and the emission filter is integrally integrated with the environmental control room.

30. The system according to claim 1, wherein, The biological sample includes cells.

31. The system according to claim 30, wherein, The cells in question are cardiomyocytes.

32. The system of claim 1, further comprising a controller that operates the irradiation module, the detection module, and, if present, the stage.

33. The system according to claim 32, wherein, The controller further operates the environmental control room.

34. The system according to claim 32, wherein, The controller instructs the detection module to detect the response light at a series of times as part of a kinetic measurement.

35. The system according to claim 32, wherein, The controller instructs the detection module to detect the response light at discrete times as part of the endpoint determination.

36. The system of claim 1, further comprising a biological sample.

37. The system according to claim 36, wherein, The biological sample includes cells.

38. The system according to claim 37, wherein, The cells in question are cardiomyocytes.

39. The system according to claim 1, wherein, The illumination light is fluorescent excitation light, and the response light is fluorescent emission light, further including a fluorophore that reports the state of the biological sample.

40. The system according to claim 39, wherein, The fluorophore reports the activity of ion channels in the biological sample.

41. The system according to claim 40, wherein, The ion is calcium.

42. The system according to claim 1, wherein, The irradiation module is further configured to generate heating light, which is used to maintain or increase the temperature of the biological sample when the biological sample is positioned at the examination point, and wherein the intensity of the heating light and the intensity of the irradiation light can be changed independently.

43. The system according to claim 42, wherein, The heating light is infrared radiation.

44. The system of claim 1, further comprising a temperature sensing element located at the inspection point.

45. The system according to claim 1, wherein, The irradiation module includes at least one LED, solid-state light source, or laser.

46. ​​The system according to claim 1, wherein, The illumination module includes multiple light sources, and the intensity of the light produced by each light source can be independently controlled to allow for changes in the spectral composition of the illumination light.

47. The system according to claim 1, wherein, The detection module includes a point detector array.

48. The system according to claim 1, wherein, The detection module includes a camera.

49. The system according to claim 1, wherein, The sampling rate of the detection module is greater than approximately 100 Hz.

50. A method for examining a biological sample, comprising: Select biological samples; Choose the system according to claim 1; The biological sample was irradiated with illumination light; Detecting the response light transmitted from the biological sample in response to the irradiation light; and The state of the biological sample is determined using the response light.

51. An environmental control room, comprising: An enclosed volume having at least one pair of substantially opposing walls, each of which is at least partially transparent; as well as A control mechanism configured to allow maintaining or altering the environmental conditions within the room.

52. The environmental control room according to claim 51, wherein, The spectrum of light that can be transmitted through one of two opposing walls is different from the spectrum of light that can be transmitted through the other of the opposing walls.

53. The environmental control room according to claim 51, wherein, At least one of the opposing walls further includes at least one lens for changing the focal point of light passing through the wall.

54. The environmental control room according to claim 51, wherein, The at least one lens is a converging lens.

55. The environmental control room according to claim 51, wherein, The at least one lens is a Fresnel lens.

56. The environmental control room according to claim 53, wherein, The at least one lens is a regular array of lenses, each lens focusing light to a lateral position of the regular array in the sample plane and / or collecting light from a lateral position of the regular array in the sample plane.

57. The environmental control room according to claim 51, wherein, The control mechanism includes a partially transparent coating disposed on at least one of the opposing walls, which heats up when an electric current passes through the coating.

58. The environmental control room according to claim 57, wherein, The coating comprises indium tin oxide.

59. The environmental control room according to claim 57, wherein, The control mechanism further includes electrical contacts for guiding electricity through the conductive coating.

60. The environmental control room according to claim 51, wherein, The control mechanism includes at least one port configured to receive a gas input.

61. The environmental control room according to claim 60, wherein, The gas is carbon dioxide.

62. The environmental control room according to claim 52, wherein, At least one of the opposing walls includes a spectral filter configured to alter the spectrum of light passing through it.

63. The environmental control room according to claim 63, wherein, Each of the opposing walls includes a spectral filter, wherein the spectrum of light passing through each spectral filter is different from the spectrum of light passing through the other spectral filter.

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