Systems for examining biological samples under controlled environmental conditions

Abstract

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

Description

SYSTEMS FOR EXAMINING BIOLOGICAL SAMPLES UNDER CONTROLLED ENVIRONMENTAL CONDITIONSCross-References to Related Applications

[0001] This application is based upon and claims the benefit under 35 U.S.C. § 119(e) of U.S. Provisional Patent Application Serial No. 63 / 583,580, filed September 18, 2023. This application incorporates by reference in their entireties for all purposes U.S. Provisional Patent Application Serial No. 63 / 583,580, filed September 18, 2023, U.S. Patent No. 11 ,041 ,806, issued June 22, 2021 , and U.S. Patent Application Publication No. US-2016-0123886-A1 , published May 5, 2016. The systems described herein may be used in conjunction with systems, features, and assays described in the incorporated patent documents.Introduction

[0002] Systems for analysis of biological samples are useful in medical research, life sciences, and other applications. Existing systems to measure the properties of such samples, alone or when combined with an effector such as a candidate drug compound, commonly include an array of wells, such as a microplate, with cells and supporting materials added to each well. The wells may then be analyzed serially or in parallel, depending on both assay design and instrumentation. Significantly, many such assays are performed under nonphysiological conditions, such as at room temperature instead of physiological temperature, potentially affecting the results. Thus, there is a need for systems for examining biological samples under controlled environmental conditions.Summary

[0003] The present disclosure provides systems, including apparatus and methods, for examining biological samples under controlled environmental conditions.Brief Description of The Drawings

[0004] FIG. 1 is a high-level schematic view of an exemplary system for examining a biological sample under controlled environmental conditions, in accordance with aspects of the present disclosure. The schematic discloses threealternative positions of an environmental control chamber (ECC): (A) external, (B) internal in a non-examination area, and (C) internal in an examination area.

[0005] FIG. 2 is a pair of isometric views of an embodiment of the system of FIG. 1 , minus the environmental control chamber and portions of a housing, showing(A) a view from above a front corner of the system, and (B) a view from above an opposite rear corner of the system, highlighting an illumination module, a detection module, a controller, and a housing.

[0006] FIG. 3 is a top view of the system of FIG. 2.

[0007] FIG. 4 is a side elevation view of the system of FIG. 2.

[0008] FIG. 5 is a schematic side view of a first exemplary system for examining a biological sample under controlled environmental conditions, in which the system has an external environmental control chamber, showing configurations (A) before,(B) during, and (C) after sample examination. This system corresponds to alternative (A) in FIG. 1.

[0009] FIG. 6 is a schematic side view of a second exemplary system for examining a biological sample under controlled environmental conditions, in which the system has an internal environmental control chamber disposed at a location distinct from where samples are examined, showing configurations (A) before, (B) during, and(C) after examination. This system corresponds to alternative (B) in FIG. 1 .

[0010] FIG. 7 is a schematic side view of a third exemplary system exemplary system for examining a biological sample under controlled environmental conditions, in which the system has an internal environmental control chamber disposed at a location where samples are examined, showing configurations (A) before and during examination, and (B) after examination. This system corresponds to alternative (C) in FIG. 1.

[0011] FIG. 8 is a schematic side view of an environmental control chamber with integrated optics.

[0012] FIG. 9 is a schematic side view of an environmental control chamber with integrated glass heaters.

[0013] FIG. 10 is schematic side view of an environmental control chamber with integrated optics and glass heaters.

[0014] FIG. 11 is a partially sectional side elevation view of the detection module of the system of FIGS. 2-4 including the environmental control chamber of FIG. 10.Detailed Description

[0015] The present disclosure provides systems, including devices and methods, for examining biological samples under controlled environmental conditions. The control may be exerted before, during, and / or after examination. The systems may include an illumination module, a detection module, and an environmental control chamber. The illumination module may be configured to produce illumination and to direct the illumination light to an examination site. The detection module may be configured to detect response light produced by one or more biological samples positioned at the examination site in response to illumination by the illumination light. The environmental control chamber may be configured to control an environmental condition of the biological sample before, during, and / or after its examination (in the lattermost case, prior to further examination, among others).

[0016] FIG. 1 is a high-level schematic view of an exemplary system 20 for examining a biological sample with environmental control, in accordance with aspects of the present disclosure. The system includes an illumination module 22, a detection module 24, and an environmental control chamber (EEC) 26. The biological sample 28, which may include one or more cells 29, may be positioned in a suitable sample holder 30. Some or all of the system may be located in a housing 31. The sample holder and sample may be positioned at an examination site 32 for analysis. Illumination light 34 produced by the illumination module may be directed onto the biological sample, when it is located at the examination site, and response light 36 from the biological sample may be directed onto (or into) the detection module. The environmental control chamber is shown in three alternative positions relative to the housing: (A) external, (B) internal, in a non-examination area, and (C) internal, in an examination area. The sample holder, and the biological sample(s) it contains, may be shuttled by a carriage or other mechanism between the external (A) or internal (at a non-examination area) (B) environmental control chambers and the examination site 32 as shown by the double-headed horizontal arrows ( ^) a and b, respectively. A controller 38 may operate or otherwise control one or more components of the system.

[0017] The illumination module may be used to generate illumination light capable of monitoring a status and / or eliciting a response from the sample. The module may include one or more individual light sources (e.g., one, two, three, four, five, six, seven, eight, nine, or more sources). The light sources may include lasers,light pipes, and / or light-emitting diodes (LEDs), among others. The lasers may include single- and / or multi-mode lasers. Each light source may be capable of emitting at one or more predominantly single wavelengths (e.g., 488 nm or 514 nm) or over one or more ranges of wavelengths (e.g., 450 nm to 550 nm). In some cases, two or more light sources may output light having the same spectral qualities, where the light from the two or more sources is combined to increase its intensity. In other cases, two or more light sources may output light having different spectral qualities, expanding the range of available illumination wavelengths such that the light engine can be used with a broader range and number of samples. The number of light sources (or sets of light sources) may be independent of or proportional to the number of samples and / or sample sites being examined. For example, there may be one light source (or set of light sources) per sample or sample site, one light source (or set of light sources) for every two samples or sample sites, one light source (or set of light sources) for every four samples or sample sites, etc. The intensity of light from each light source may be independently adjustable, for example, from 0% to 100% relative intensity. Light output by the illumination module during a particular analysis or assay may come from a single source or be a blend of light from two or more sources. The spectral properties of light output by the light engine may be matched to its intended use, for example, to activate a photoexcitable compound and / or process, and / or to excite fluorescence from preselected fluorescent tags, among others. Multiple wavelengths of light may be applied either simultaneously or sequentially, according to the requirements of the specific activation and / or examination protocol. The illumination module may optionally include a diffuser, despeckler, and / or other mechanism for reducing inhomogeneities in the illumination light. The illumination module may include reflective and / or refractive elements for combining light from different light sources onto a single optical pathway. The illumination light, in use, may illuminate all or part of the biological sample(s). When the sample is housed in a multi-well plate, the illumination light may simultaneously illuminate one, some, many, or all of the wells of the plate. The illumination light may be particularly suitable for the photostimulation of cellular responses and the recording of cellular responses. The illumination module may include, and / or direct 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 may be used to capture response light generated by the biological sample in response to illumination by the illumination light and toproduce a corresponding signal (or signals) or other representation of the response light for further analysis. The biological sample and / or array of biological samples may have a nonzero spatial extent transverse to the illumination direction and / or comprise a series of discrete samples again having a nonzero spatial extent. Consequently, the detection module may be capable of detecting response light over 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 in 1 :1 spatial registration with wells of a multi-well sample holder, with one or more lenses or lens arrays used to focus response light from each well onto its respective detector. Alternatively, or in addition, the detection module may include an imaging detector, such as a camera, including a complementary metal-oxide semiconductor (CMOS) camera, a scientific-CMOS (sCMOS) camera, and / or a charge-coupled device (CCD) camera, among others. Signals from multiple points in the sample, or from multiple samples, may be collected simultaneously by all elements of an array detector or all portions of an imaging detector (i.e., by parallel sampling). Signal acquisition may be rapid (e.g., from about 10 Hz to about 10 kHz, among others). High sampling rates provide the temporal resolution needed for accurate tracking of cellular responses on millisecond timescales.

[0019] The environmental control chamber may be used to control one or more environmental conditions of the biological sample, such as temperature, gas composition, humidity, and / or pressure, among others. These conditions may be held substantially constant. For example, temperature may be held at or near a physiological temperature, such as 37°C. Alternatively, or in addition, conditions may be varied (e.g., ramped, cycled, or otherwise varied between different values). For example, temperature may be increased or decreased from a base value to assess the effect(s) of heating or cooling on a sample. The environmental control chamber may include a port, such as a lid and associated opening, for introducing and removing a sample holder, and associated biological samples, from the chamber. The environmental control chamber may further include sensors that monitor one or more environmental conditions within the chamber and effectors to bring about a desired change in the monitored condition(s). 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 effectormay be integral to the chamber, such as a heating or cooling slab, or external, such as an infrared light source in the illumination module that produces infrared light for absorption by the sample and / or associated sample holder. In the latter case, a filter may be used between the sample(s) and detector to reduce or eliminate the infrared radiation before it can be detected and confused with response light. The chamber may further include, or be associated with, optics for focusing or otherwise directing illumination light onto the biological sample(s) and collecting response light generated by the sample(s) and directing it to the detection module.

[0020] The biological sample may include any particle(s), substance(s), extract(s), mixture(s), and / or assembly(ies) derived from or corresponding to one or more organisms, cells, and / or viruses. The biological sample also may include additional components to facilitate analysis, such as fluid (e.g., water), buffer, culture nutrients, salts, other reagents, dyes, etc. The biological sample will commonly include cells. These cells may be disposed in any suitable medium, such as a culture medium. In some cases, the cells may be adherent cells that are cultured preferentially or exclusively attached to, and / or in contact with, a substrate, such as glass or plastic. Suitable cells may include cells derived from animals, plants, bacteria, and / or fungi (including yeast), among others. The cells may be primary, immortalized, senescent, stem, differentiated, transformed, infected, transgenic, assembled into tissues or multi-cellular organisms (such as embryos), and / or the like. Substances, as used here, 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 noncyclic mononucleotides, DNA, and / or RNA), carbohydrates, lipids, ions, hormones, metabolites, conjugates thereof, and / or the like. Extracts and / or assemblies, as used here, may include fractions or organelles isolated from cells. Exemplary fractions and / or organelles may include nuclei, membranes, mitochondria, cytosol, whole cell lysates, etc., either crude or at least partially purified. Assemblies also may include 3D spheroids. The sample may be selected to facilitate the screening of compounds, such as candidate pharmaceuticals, for biological activity (including a lack of (harmful) activity during safety screening). For example, the sample may be selected to facilitate the rapid screening of pharmacologically active compounds against ion-channel targets or safety pharmacology assays. Exemplary cells include cardiomyocytes, among others. For simplicity, the term “sample,” as used in the disclosure, may referto a single sample or multiple samples, depending on the context. For example, the expression “sample holder and sample” may refer to a single sample if the sample holder is configured to hold a single sample or multiple samples if the sample holder, such as a multi-well plate, is configured to hold (and is loaded with) multiple samples.

[0021] The biological sample may be supported or otherwise held by a suitable sample holder. The sample holder generally comprises any substrate or other mechanism for holding samples for illumination and detection. The sample holder may hold one or more discrete samples at one or more distinct sample sites. In some cases, sample sites may be defined by mechanical barriers, such as walls, for example, forming sample wells. In other cases, sample sites may be defined by chemical barriers, such as hydrophobic regions separating hydrophilic regions, or distinct spatially separated binding sites for nucleic acids, proteins, and / or other materials. The sample sites may be separate fluid volumes or share a common fluid volume. Exemplary sample holders with separate fluid volumes may include multi-well plates, such as microplates and PCR plates, among others. Such plates may have any suitable number of sample wells, such as 4, 6, 12, 24, 48, 96, 384, or 1536 sample wells, among others. Exemplary sample holders with a common fluid volume may include microscope slides and / or sample chips for nucleic acids and / or proteins, among others. The samples in all cases may be independent of one another or aliquots or replicates of one another, depending on the analysis. The samples may include control or calibration samples. The sample holder may be at least partially transparent to illumination and detection light, at least in the predominant direction of light propagation during examination.

[0022] The carriage may include any structure configured to support and move the sample holder and associated biological samples for input, output, and / or examination. It may allow samples to be added to and removed from the system (e.g., at an input / output region). It also may convey samples between the environmental control chamber and examination site when the two are not coextensive. The carriage may include rails, belts, gears, pulleys, and motors, among others, with its motion directed by the controller.

[0023] The controller may include any mechanism for assessing the state of the system (e.g., the location, identity, and / or status of a sample), for running the optics (e.g., determining and executing an illumination protocol and / or collecting the resulting response light), for conveying the sample holder and sample between an input / outregion, the environmental control chamber, and the examination site (if different). The controller also may monitor and report on the condition of the system itself and its subcomponents.

[0024] The system further may include a variety of optical elements, or optics, for performing a corresponding variety of purposes. These optics may be directly associated with one or more major system components, such as the illumination module, the detection module, and the environmental control chamber, among others. These optics also may be dispersed, as appropriate, between system components. Examples of optical elements include mirrors, lenses, beamsplitters, spectral filters (such as fluorescence excitation and emission filters), and neutral density filters, among others. The optics may act on the illumination light, the response light, and / or any additional light (such as infrared radiation used to heat the sample). Mirrors use reflection to alter a direction and / or focus of the light, allowing among other uses light from different sources to be combined and the optical path through the system to be folded, reducing its footprint. Lenses use refraction to bend light, allowing the light to be focused or collimated or decollimated. This is especially important where illumination light is directed onto the sample(s) and response light is collected from the sample and directed onto the detector(s). Beamsplitters separate light, typically transmitting some light along one path and reflecting the rest along another path. The separation may or may not depend on the spectrum of the light (e.g., in the former case, reflecting shorter wavelengths and passing longer wavelengths). Spectral filters partially or completely block light with some wavelengths and partially or completely transmit light with other wavelengths. These filters may play an important role in “cleaning up” light from the illumination module (e.g., by reducing or eliminating light with undesired wavelengths before it impinges on a sample). They also may play an important role in fluorescence assays. A filter placed before the sample, known as an excitation filter, can reduce or remove wavelengths of light that are not needed to excite fluorescence (particularly if those wavelengths could be confused with fluorescence emission light). A filter placed after the sample, known as an emission filter, can reduce or remove wavelengths of light not associated with fluorescence emission (particularly wavelengths corresponding to stray excitation light), helping to ensure that the detected light is response light by reducing contamination. Emission filters are especially important when excitation light and collected emission light follow the same optical path (i.e., in trans- versus epi-fluorescence configurations). Neutraldensity filters may reduce the intensity of light, including illumination light and / or response light, at least substantially uniformly (neutrally) across the wavelengths of interest. This may improve performance if the intensity is otherwise too bright. Alternatively, or in addition, the intensity of the illumination light (and thus indirectly the response light) can be controlled by the illumination module itself, for example, by altering the strength and / or duration of power supplied to the light sources. The relative choices, positions, sizes, and / or other attributes of these various optics may be selected to increase or decrease the portion of the sample holder (and thus the sample) that is illuminated and / or to alter the quality of that illumination.

[0025] Lenses may play a particularly significant role in the system and may be positioned in the path taken by illumination light, the path taken by response light, or both paths. These lenses may perform any suitable function. For example, a lens positioned in the illumination path may homogenize and collimate illumination light incident on the sample holder, such that its intensity is more uniform and / or is more nearly parallel to the optical axis and / or perpendicular to a plane of the sample holder, reducing shadows. Alternatively, or in addition, a lens positioned in the response path may collect response light and direct it toward the detection module, increasing the amount of light captured by the detection module. The lens also may focus light onto the detection module, ensuring that the position of light on the detector is better correlated with its position of origin in the sample. Lenses disposed between major components of the system may complement or supplement the role of lenses integral with the illumination module, detection module, and / or environmental control chamber. Lenses may variously and without limitation be termed condenser lenses, objective lenses, field lenses, and / or tube lenses, among others, depending on their positions and functions. The lenses also may work in pairs or triplets, among others, along a given path, and / or as arrays of lenses, among others, transverse to the illumination and response paths.

[0026] FIGS. 2-4 show various views of portions of an embodiment 50 of the system of FIG. 1. The portions include an illumination module 52, a detection module 54, a controller 56, and a housing 58. The three possible locations A, B, C of the environmental control chamber portrayed in FIG. 1 are shown and described in FIGS. 5-7, respectively, and described in the associated text.

[0027] FIG. 5 is a schematic side view of a first exemplary system 70 for examining a biological sample. Here, an environmental control chamber 72 is locatedexternal to a housing 74 containing other components of the system used to conduct the examination. FIG. 5 shows three exemplary configurations associated with sample examination. (A) Before examination, a sample holder 76 (containing a biological sample is positioned within the environmental control chamber and kept there until examination occurs. A temperature or other environmental condition is imposed and / or altered. The sample is then moved into position for examination by opening a door 78 separating the environmental control chamber and housing and conveying the sample holder and sample through the door to the examination site using a carriage 80. (B) During examination, the door between the environmental control chamber and housing is closed, the sample holder is positioned at the examination site, the sample is illuminated by illumination light, and the resulting response light is detected. (C) After examination, the door between the housing and environmental control chamber is reopened, and the sample holder and sample are conveyed back to the environmental control chamber. The same sample may be analyzed again by repeating the steps associated with configurations A and B. Alternatively, as shown, a lid 82 or other closure mechanism for the environmental control chamber may be opened and the sample holder and sample removed. The entire process may be repeated for a new sample. The environmental control chamber here and elsewhere in the disclosure may house a single sample holder or multiple sample holders, depending on the embodiment. In the latter case, the carriage will select the sample holder and sample of interest based on instructions from the controller.

[0028] FIG. 6 is a schematic side view of a second exemplary system 90 for examining a biological sample. Here, an environmental control chamber 92 is located inside a housing 94 containing other components of the system used to conduct the examination. However, the location of the environmental control chamber is distinct and different from the examination site 96 (dashed rectangle) where the sample is examined. FIG. 6 shows three exemplary configurations associated with sample examination. (A) Before examination, a sample holder 98 containing a biological sample is positioned within the environmental control chamber. A temperature or other environmental condition is imposed and / or altered. The sample is then moved into the examination site for examination by moving the sample holder using a carriage 100. (B) During examination, the sample is illuminated by illumination light, and the resulting response light is detected. (C) After examination, a door 102 in the housing is opened, and the sample holder is conveyed outside by the carriage and removed.The same sample may 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 configuration C have been completed, a new sample holder may be placed on the carriage, the sample holder may be conveyed to the environmental control chamber, the door in the housing may be closed, and the new sample may be examined as described above. Removing the sample from the environmental control chamber during examination may decrease the complexity of the chamber.

[0029] FIG. 7 is a schematic side view of a third exemplary system 110 for examining a biological sample. Here, like in FIG. 6, an environmental control chamber 112 is located inside a housing 114 containing other components of the system used to conduct the examination. However, in this embodiment, the examination site 116 (dashed rectangle) is located inside rather than outside the environmental control chamber. Thus, samples may be examined while the samples are inside the environmental control chamber, ensuring better control of sample condition(s), while increasing the complexity of the environment control chamber (which must now be at least partially transparent). FIG. 7 shows two exemplary configurations associated with sample examination. (A) Before and during examination, a sample holder 118 is positioned at the examination site in the environmental control chamber. A temperature or other environmental condition is imposed and / or altered. The sample is illuminated by illumination light, and the resulting response light is detected. The sample may be interrogated at a single time or time interval, or at distinct 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 conveyed outside using a carriage 122 and removed. A new sample holder may then be placed on the carriage, the new sample holder conveyed into the environmental control chamber, the door in the housing closed, and the new sample can be examined as described above. Suitable environmental control chambers for this embodiment are shown in FIGS. 8-10.

[0030] Exemplary details of suitable illumination modules, detection modules, controllers, and housings for the embodiments in FIGS. 5-7 are shown in FIGS. 2-4, among other places in the present disclosure, and are further described in the cross- referenced patent documents.

[0031] FIG. 8 is a schematic side view of an environmental control chamber 130 with integrated optics. The control chamber includes a central region 132 having a controllable environment and sized to receive at least one sample holder. The central region is disposed between two at least substantially planar, at least substantially parallel, walls. A first, illumination wall 134 is configured to transmit illumination light 135 from an illumination module toward a sample holder and associated biological sample(s) contained within the chamber. A second, detection wall 136 is configured to transmit response light 137 produced by the biological sample to a detection module. Here, the illumination and detection walls may be formed using any material capable of transmitting all or a usable part of the illumination light and detection light. Suitable materials may include clear plastic and / or glass. The walls, in turn, may include suitable illumination and detection optics 138a,b, such as lens arrays. The number of lenses, or lenslets, in the lens array may be matched to a number of samples, or sample wells, in the sample holder, such that each lens illuminates (138a) and / or detects from (138b) a single sample or well. The optics may be molded into the plastics.

[0032] FIG. 9 is a schematic side view of an environmental control chamber 150 with integrated glass heaters. Environmental control chamber 150 shares many similarities with environmental control chamber 130 shown in FIG. 8. For example, like control chamber 130, control chamber 150 includes a central region 152 having a controllable environment and sized to receive at least one sample holder. Moreover, the central region is disposed between two at least substantially planar, at least substantially parallel, walls, namely, an illumination wall 154 configured to transmit illumination light 155 and a detection wall 156 configured to transmit detection light 157. However, in the present embodiment, one or both of the walls includes an optically transparent heating element 158a, b. The walls may comprise clear glass, and the heating element may comprise an indium tin oxide (and / or other electrically heatable) coating. If the coating is deposited in a circuitous path (e.g., in a serpentine path and / or a zigzag path, among others) and a voltage difference is applied between one end of the path and the other, then resistive heating will occur. This heating can be used as a thermal input to the environmental chamber, allowing a temperature to be established and / or maintained. Moreover, because indium tin oxide can reflect infrared light, it may be used as both a filter and a heater.

[0033] FIG. 10 is schematic side view of an environmental control chamber 170 with integrated optics (like FIG. 8) and glass heaters (like FIG. 9). Environmental control chamber 170 again includes a central region 172 having a controllable environment, sized to receive at least one sample holder, and at least substantially planar parallel illumination and detection walls 174, 176, as defined above. Moreover, one or both walls include both integrated optics and integrated heating elements, as described above. The integrated features on the upstream (illumination or excitation) side of the environmental control chamber may include one or more of illumination light collimating optics 174a, illumination light filter(s) 174b, an illumination light focusing lens element array 174c, and illumination light side heating element, among others. These integrated features collimate 174a incoming illumination light 175, filter 174b the light to remove unintended or undesirable wavelength bands and / or reduce intensity, focus 174c the light on the sample, and control the temperature 174d in the sample chamber. The integrated features on the downstream (detection or emission) side of the environmental control chamber may include one or more of detection light collimating optics 176a (such as an emission light collimating lens element array), detection light filter(s) 176b, a detection light focusing lens element array 176c, and a detection side heating element 176d, among others. These integrated features collimate 174a the response light 177, filter 174b it, and then refocus 174c the filtered light onto the sensor or sensor array in the detection module, while again controlling the temperature 176d of the sample chamber. Aspects of the environmental control chamber, including the various optics and heating elements, may be disposed in any suitable order, although the heating elements typically will be disposed closest to the sample holder.

[0034] FIG. 11 is a partially sectional side elevation view of the detection module (or reading head) 54 of FIGS. 2-4 (including the environmental control chamber 170 of FIG. 10). The light path is generally indicated by the arrows showing excitation (illumination) light 185 and emission (detection) light 187. The figure shows the detection module in use, with a multi-well sample holder 190 and associated sample wells / samples 191 disposed for examination within an environmental control chamber 192. An upper heating element 194a and a lower heating element 194b are disposed above and below the sample holder on the upstream excitation (illumination) side and downstream emission (detection) side of the examination site and sample holder. The heating elements may be used to maintain and / or alter the temperature ofthe sample holder and associated sample(s). Excitation light 185 from an illumination module is directed toward the examination site, here by a pair of 100% mirrors 196a,b. 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 sites (e.g., sample wells) in the sample holder, an excitation filter 202, which cleans up the excitation light, and the upper heating element. Emission (e.g., fluorescence) light 187 emitted from the sample passes through the lower heating element, an emission objective lens array 204, which collects and optionally collimates the emission light, an emission filter 206, which cleans up the emission light (in particular, removing contaminating excitation light), an emission tube lens array 208, which focuses the light onto a detector (e.g., a detection printed circuit board assembly (PCBA)) 210, which detects the light and converts it into an electronic signal for processing by the controller. A carriage 212 shuttles the sample holder and associated sample(s) to and from the examination site.Selected Aspects

[0035] This section describes additional selected aspects of the present disclosure, presented without limitation as a series of paragraphs, some or all of which may be numerically indexed for clarity and efficiency. Each of these paragraphs can be combined with one or more other paragraphs, and / or with disclosure from elsewhere in this application, in any suitable manner. Some of the paragraphs below expressly refer to and further limit other paragraphs, providing without limitation examples of some of the suitable combinations.1. A system for examining a biological sample, comprising (A) an illumination module configured to produce illumination light and to direct the illumination light to an examination site; (B) a detection module configured to detect response light produced by a biological sample positioned at the examination site in response to illumination by the illumination light; and (C) an environmental control chamber configured to control an environmental condition of the biological sample before and / or during its examination.2. The system of paragraph 1 , further comprising a housing defining an inside and an outside, wherein the illumination module, detection module, and examination site are disposed at least substantially inside the housing.3. The system of paragraph 2, wherein the environmental control chamber is positioned outside the housing.4. The system of paragraph 3, further comprising a carriage that transports biological samples between the environmental control chamber and the examination site.5. The system of paragraph 2, wherein the environmental control chamber is positioned inside the housing.6. The system of paragraph 5, wherein the environmental control chamber is positioned outside the examination site, further comprising a carriage that transports biological samples between the environmental control chamber and the examination site.7. The system of paragraph 5, wherein the environmental control chamber is positioned at the examination site.8. The system of paragraph 7, the environmental control chamber having an at least partially transparent illumination side through which illumination light enters the environmental control chamber from the illumination module and an at least partially transparent detection side through which response light exits the environmental control chamber toward the detection module, wherein the illumination side and the detection side are at least substantially planar and parallel to one another.9. The system of paragraph 8, wherein at least one of the illumination side and the detection side includes a heating element.10. The system of paragraph 9, wherein the heating element is at least partially transparent and disposed in the path of the illumination light, response light, or both.11. The system of paragraph 10, wherein the heating element includes an indium tin oxide coating.12. The system of any of paragraphs 7 to 11 , wherein the environmental control chamber includes at least one lens disposed on one or both of the illumination side and the detection side.13. The system of paragraph 12, wherein the biological sample is housed in a multi-well plate, and wherein the environmental control chamber includes a lens for each well in the multi-well plate.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 for each well, and one lens on the detection side for each well.15. The system of paragraph 14, wherein the environmental control chamber further includes at least three lenses for each well in the multi-well plate, two lenses on one side of the multi-well plate for each well, and at least one lens on the other side of the multi-well plate for each well.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 for each well, and two lenses on the detection side for each well.17. The system of any of paragraphs 13 to 16, wherein the environmental control chamber includes at least one Fresnel lens on the illumination side.18. The system of any of paragraphs 13 to 17, wherein the detection module collects response light separately from each well, so that an intensity of light from each well can be distinguished from an intensity of light from each other well.19. The system of any of paragraphs 13 to 18, wherein the number of wells in the multi-well plate is selected from the group consisting of 4, 6, 12, 24, 48, 96, 384, and 1536.20. The system of any of paragraphs 12 to 19, wherein the at least one lens is at least one Fresnel lens.21. The system of any preceding paragraph, wherein the environmental condition is selected from the group consisting of a temperature and an atmospheric component.22. The system of paragraph 21 , wherein the environmental condition is a temperature.23. The system of paragraph 21 , wherein the environmental condition is a carbon dioxide concentration.24. The system of any preceding paragraph, wherein the illumination light is fluorescence excitation light, and wherein the response light is fluorescence emission light.25. The system of paragraph 24, wherein the detection module determines an intensity of the fluorescence emission light.26. The system of paragraph 25, the biological sample consisting of at least two discrete samples, wherein the detection module determines an intensity of the fluorescence emission light for each discrete sample.27. The system of any of paragraphs 24 to 26, further comprising an excitation filter operatively disposed between the illumination module and the examination site and configured to alter a spectral property of the illumination light before it is incident on the biological sample.28. The system of any of paragraphs 24 to 27, further comprising an emission filter operatively disposed between the examination site and the detection module and configured to alter a spectral property of the response light before it is incident on the detector.29. The system of paragraph 27 or 28, wherein the environmental control chamber is positioned at the examination site, and wherein at least one of the excitation filter and the emission filter is integral with the environmental control chamber.30. The system of any preceding paragraph, wherein the biological sample includes cells.31 . The system of paragraph 30, wherein the cells are cardiomyocytes.32. The system of any preceding paragraph, further comprising a controller that operates the illumination module, the detection module, and, if present, the carriage.33. The system of paragraph 32, wherein the controller further operates the environmental control chamber.34. The system of paragraph 32 or 33, wherein the controller instructs the detection module to detect response light at a series of times as part of a kinetic assay.35. The system of paragraph 32 or 33, wherein the controller instructs the detection module to detect response light at a discrete time as part of an endpoint assay.36. The system of any previous paragraph, further comprising a biological sample.37. The system of paragraph 36, wherein the biological sample includes cells.38. The system of paragraph 37, wherein the cells are cardiomyocytes.39. The system of any preceding paragraph, wherein the illumination light is fluorescence excitation light, and wherein the response light is fluorescence emission light, further comprising a fluorophore that reports on a status of the biological sample.40. The system of paragraph 39, wherein the fluorophore reports on the activity of an ion channel in the biological sample.41 . The system of paragraph 40, wherein the ion is calcium.42. The system of any preceding paragraph, wherein the illumination module is further configured to produce heating light used to maintain or increase a temperature of the biological sample when it is positioned at the examination site, and wherein an intensity of the heating light and an intensity of the illumination light can be independently varied.43. The system of paragraph 42, wherein the heating light is infrared radiation.44. The system of any preceding paragraph, further comprising a temperature sensing element positioned at the examination site.45. The system of any preceding paragraph, wherein the illumination module includes at least one LED, solid-state light source, or laser.46. The system of any preceding paragraph, wherein the illumination module includes a plurality of light sources, and wherein the intensity of light produced by each light source can be independently controlled, allowing the spectral content of the illumination light to be varied.47. The system of any preceding paragraph, wherein the detection module includes an array of point detectors.48. The system of any preceding paragraph, wherein the detection module includes a camera.49. The system of any preceding paragraph, wherein a sampling rate of the detection module is greater than about 100 Hertz (Hz).50. A method of examining a biological sample, comprising (A) selecting a biological sample; (B) selecting a system according to any preceding paragraph; (C) illuminating the biological sample using illumination light; (D) detecting response light transmitted from the biological sample in response to the illumination light; and (E) determining a status of the biological sample using the response light.51 . An environmental control chamber, comprising (A) an enclosed volume having at least one pair of generally opposed walls, each of the opposed walls being at least partially transparent; and (B) a control mechanism configured to allow maintenance or alteration of an environmental condition within the chamber.52. The environmental control chamber of paragraph 51 , wherein the spectrum of light that can be transmitted through one of the two opposed walls is different from the spectrum of light that can be transmitted through the other of the opposed walls.53. The environmental control chamber of paragraphs 51 or 52, wherein at least one of the opposed walls further includes at least one lens for altering the focus of light passing through the walls.54. The environmental control chamber of any of paragraphs 51 to 53, wherein the at least one lens is a converging lens.55. The environmental control chamber of any of paragraphs 51 to 54, wherein the at least one lens is a Fresnel lens.56. The environmental control chamber of any of paragraphs 53 to 54, wherein the at least one lens is a regular array of lenses, each focusing light to a regular array of lateral positions in a sample plane and / or collecting light from a regular array of lateral positions in a sample plane.57. The environmental control chamber of any of paragraphs 51 to 56, wherein the control mechanism includes an at least partially transparent coating disposed on at least of the opposed walls that heats up when an electric current is passed through the coating.58. The environmental control chamber of paragraph 57, wherein the coating includes indium tin oxide.59. The environmental control chamber of paragraph 57 or 58, wherein the control mechanism further includes electric contacts for directing electricity through the conductive coating.60. The environmental control chamber of any of paragraphs 51 to 59, wherein the control mechanism includes at least one port configured to receive a gas input.61 . The environmental control chamber of paragraph 60, wherein the gas is carbon dioxide.62. The environmental control chamber of any of paragraphs 51 to 61 , wherein at least one of the opposed walls includes a spectral filter configured to alter a spectrum of light passing through the filter.63. The environmental control chamber of paragraph 62, wherein each of the opposed walls includes a spectral filter, and wherein a spectrum of light passed by each spectral filter differs from a spectrum of light passed by the other spectral filter.

[0036] The term “and / or” as used in the present disclosure means all combinations of the listed elements. For example, a list with two elements “A and / or B” covers three possibilities: only A, only B, and both (A and B). Similarly, a list with three elements “A, B, and / or C” covers seven possibilities: only A, only B, only C, both A and B, both A and C, both B and C, and all three (A, B, and C). The extension to four or more elements follows the same pattern.

[0037] The term "exemplary" as used in the present disclosure means "illustrative" or "serving as an example" and is not intended to imply desirability or superiority.

[0038] The terms “first,” “second,” “A,” “B,” and so on are used in the present disclosure to distinguish or identify various members of a group, or the like, in the order in which they are introduced in a particular context and are not intended to show serial or numerical limitation.

[0039] The disclosure set forth above may encompass multiple distinct inventions with independent utility. Although each of these inventions has been disclosed in its preferred form(s), the specific embodiments thereof as disclosed and illustrated herein are not to be considered in a limiting sense, because numerous variations are possible. The subject matter of the inventions includes all novel and nonobvious combinations and subcombinations of the various elements, features, functions, and / or properties disclosed herein. The following claims particularly point out certain combinations and subcombinations regarded as novel and nonobvious. Inventions embodied in other combinations and subcombinations of features, functions, elements, and / or properties may be claimed in applications claiming priority from this or a related application. Such claims, whether directed to a different invention or to the same invention, and whether broader, narrower, equal, or different in scopeto the original claims, also are regarded as included within the subject matter of the inventions of the present disclosure.

Claims

WHAT IS CLAIMED:1 . A system for examining a biological sample, comprising: an illumination module configured to produce illumination light and to direct the illumination light to an examination site; a detection module configured to detect response light produced by a biological sample positioned at the examination site in response to illumination by the illumination light; and an environmental control chamber configured to control an environmental condition of the biological sample before and / or during its examination.

2. The system of claim 1 , further comprising a housing defining an inside and an outside, wherein the illumination module, detection module, and examination site are disposed at least substantially inside the housing.

3. The system of claim 2, wherein the environmental control chamber is positioned outside the housing.

4. The system of claim 3, further comprising a carriage that transports biological samples between the environmental control chamber and the examination site.

5. The system of claim 2, wherein the environmental control chamber is positioned inside the housing.

6. The system of claim 5, wherein the environmental control chamber is positioned outside the examination site, further comprising a carriage that transports biological samples between the environmental control chamber and the examination site.

7. The system of claim 5, wherein the environmental control chamber is positioned at the examination site.

8. The system of claim 7, the environmental control chamber having an at least partially transparent illumination side through which illumination light enters the environmental control chamber from the illumination module and an at least partially transparent detection side through which response light exits the environmental control chamber toward the detection module, wherein the illumination side and the detection side are at least substantially planar and parallel to one another.

9. The system of claim 8, wherein at least one of the illumination side and the detection side includes a heating element.

10. The system of claim 9, wherein the heating element is at least partially transparent and disposed in the path of the illumination light, response light, or both.11 . The system of claim 10, wherein the heating element includes an indium tin oxide coating.

12. The system of claim 7, wherein the environmental control chamber includes at least one lens disposed on one or both of the illumination side and the detection side.

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

14. The system of claim 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 for each well, and one lens on the detection side for each well.

15. The system of claim 14, wherein the environmental control chamber further includes at least three lenses for each well in the multi-well plate, two lenses on one side of the multi-well plate for each well, and at least one lens on the other side of the multi-well plate for each well.

16. The system of claim 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 for each well, and two lenses on the detection side for each well.

17. The system of claim 13, wherein the environmental control chamber includes at least one Fresnel lens on the illumination side.

18. The system of claim 13, wherein the detection module collects response light separately from each well, so that an intensity of light from each well can be distinguished from an intensity of light from each other well.

19. The system of claim 13, wherein the number of wells in the multi-well plate is selected from the group consisting of 4, 6, 12, 24, 48, 96, 384, and 1536.

20. The system of claim 12, wherein the at least one lens is at least one Fresnel lens.21 . The system of claim 1 , wherein the environmental condition is selected from the group consisting of a temperature and an atmospheric component.

22. The system of claim 21 , wherein the environmental condition is a temperature.

23. The system of claim 21 , wherein the environmental condition is a carbon dioxide concentration.

24. The system of claim 1 , wherein the illumination light is fluorescence excitation light, and wherein the response light is fluorescence emission light.

25. The system of claim 24, wherein the detection module determines an intensity of the fluorescence emission light.

26. The system of claim 25, the biological sample consisting of at least two discrete samples, wherein the detection module determines an 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 illumination module and the examination site and configured to alter a spectral property of the illumination light before it is incident on the biological sample.

28. The system of claim 24, further comprising an emission filter operatively disposed between the examination site and the detection module and configured to alter a spectral property of the response light before it is incident on the detector.

29. The system of claim 27, wherein the environmental control chamber is positioned at the examination site, and wherein at least one of the excitation filter and the emission filter is integral with the environmental control chamber.

30. The system of claim 1 , wherein the biological sample includes cells.31 . The system of claim 30, wherein the cells are cardiomyocytes.

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

33. The system of claim 32, wherein the controller further operates the environmental control chamber.

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

35. The system of claim 32, wherein the controller instructs the detection module to detect response light at a discrete time as part of an endpoint assay.

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

37. The system of claim 36, wherein the biological sample includes cells.

38. The system of claim 37, wherein the cells are cardiomyocytes.

39. The system of claim 1 , wherein the illumination light is fluorescence excitation light, and wherein the response light is fluorescence emission light, further comprising a fluorophore that reports on a status of the biological sample.

40. The system of claim 39, wherein the fluorophore reports on the activity of an ion channel in the biological sample.41 . The system of claim 40, wherein the ion is calcium.

42. The system of claim 1 , wherein the illumination module is further configured to produce heating light used to maintain or increase a temperature of the biological sample when it is positioned at the examination site, and wherein an intensity of the heating light and an intensity of the illumination light can be independently varied.

43. The system of claim 42, wherein the heating light is infrared radiation.

44. The system of claim 1 , further comprising a temperature sensing element positioned at the examination site.

45. The system of claim 1 , wherein the illumination module includes at least one LED, solid-state light source, or laser.

46. The system of claim 1 , wherein the illumination module includes a plurality of light sources, and wherein the intensity of light produced by each light source can be independently controlled, allowing the spectral content of the illumination light to be varied.

47. The system of claim 1, wherein the detection module includes an array of point detectors.

48. The system of claim 1 , wherein the detection module includes a camera.

49. The system of claim 1 , wherein a sampling rate of the detection module is greater than about 100 Hertz (Hz).

50. A method of examining a biological sample, comprising: selecting a biological sample; selecting a system according to claim 1 ; illuminating the biological sample using illumination light; detecting response light transmitted from the biological sample in response to the illumination light; and determining a status of the biological sample using the response light.

51. An environmental control chamber, comprising: an enclosed volume having at least one pair of generally opposed walls, each of the opposed walls being at least partially transparent; and a control mechanism configured to allow maintenance or alteration of an environmental condition within the chamber.

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

53. The environmental control chamber of claim 51 , wherein at least one of the opposed walls further includes at least one lens for altering the focus of light passing through the walls.

54. The environmental control chamber of claim 51 , wherein the at least one lens is a converging lens.

55. The environmental control chamber of claim 51 , wherein the at least one lens is a Fresnel lens.

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

57. The environmental control chamber of claim 51 , wherein the control mechanism includes an at least partially transparent coating disposed on at least of the opposed walls that heats up when an electric current is passed through the coating.

58. The environmental control chamber of claim 57, wherein the coating includes indium tin oxide.

59. The environmental control chamber of claim 57, wherein the control mechanism further includes electric contacts for directing electricity through the conductive coating.

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

61. The environmental control chamber of claim 60, wherein the gas is carbon dioxide.

62. The environmental control chamber of claim 52, wherein at least one of the opposed walls includes a spectral filter configured to alter a spectrum of light passing through the filter.

63. The environmental control chamber of claim 63, wherein each of the opposed walls includes a spectral filter, and wherein a spectrum of light passed by each spectral filter differs from a spectrum of light passed by the other spectral filter.

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