Infrared imaging system and related methods

Abstract

An infrared imaging system and a method for imaging a sample with a fluorescent marker are provided. The system includes a light source configured to illuminate a sample contact surface. The light source includes a first lighting module and a second lighting module, each configured to project a corresponding first infrared illumination beam and a second infrared illumination beam toward a sample holder, the infrared illumination beams interacting at an imaging plane to form an illumination area having a rectangular shape and uniform power distribution. The system also includes a control unit operably connected to a motor assembly and an optomechanical mechanism. The control unit is configured to cause a sampling plane and an imaging plane to overlap with each other at any of the multiple positions within the packaging shell. The system includes a detector configured to receive light emitted by the fluorescent marker of the sample when the sample is illuminated in the imaging plane when the sampling plane and the imaging plane overlap with each other.

Description

Technical Field

[0001] The technical field of the present application relates to imaging systems and related methods, and more particularly to infrared imaging systems and related methods. Background Art

[0002] Many preclinical imaging systems are commercially available. Non-limiting examples include the IVIS® Spectral In Vivo Imaging System from PerkinElmer, the Lago System from Spectral Instrument Imaging, the Pearl Trilogy® System from Li-Cor, and the Newton System from Vilber. These commercially available solutions typically rely on silicon-based detectors and, as a result, have inherent limitations in their applications because they can only image in the visible portion of the electromagnetic spectrum to the near-infrared window (NIR-1) portion of the electromagnetic spectrum (i.e., approximately 400 nm to approximately 1000 nm).

[0003] Compared to the visible or NIR-I portion of the electromagnetic spectrum, light absorption and scattering by living tissue is much weaker in the second near-infrared window (NIR-II, i.e., approximately 1000 nm to approximately 1700 nm), which means that small animals will be more transparent in the NIR-II imaging window.

[0004]

[0006] Therefore, there is a need for techniques, methods, systems, and devices that address or at least mitigate at least some of the challenges presented above. Summary of the Invention

[0005] According to one aspect, an infrared imaging system for imaging a sample having a fluorescent marker is provided. The infrared imaging system includes a housing, a sample holder, a light source, a motor assembly, an optomechanical mechanism, a control unit, and a detector. The sample holder is mounted in the housing. The sample holder has a sample contact surface and a sampling plane. The light source is configured to illuminate the sample contact surface and includes first and second illumination modules, each configured to project a corresponding first and second infrared illumination beams toward the sample holder. The first and second infrared illumination beams interact at the imaging plane to form an illumination area having a rectangular shape and a uniform power distribution. The motor assembly is configured to move the sample holder to multiple positions within the housing. The optomechanical mechanism is configured to adjust the orientation of the first and second infrared illumination beams to move the illumination area within the housing. The control unit is operably connected to the motor assembly and the optomechanical mechanism and configured to cause the sampling plane and the imaging plane to overlap at any one of the multiple positions within the housing. The detector is configured to receive light emitted by the fluorescent markers of the sample when the sample is illuminated in the imaging plane when the sampling plane and the imaging plane overlap.

[0006] In some embodiments, the encapsulating housing defines an interior volume, the encapsulating housing further comprising a door or drawer for accessing the contents of the interior volume.

[0007] In some embodiments, the sampling plane is vertically offset from the sample contact surface.

[0008] In some embodiments, the sampling plane is vertically offset from the sample contact surface by a distance value corresponding to the thickness of the sample or the thickness of a portion of the sample.

[0009] In some embodiments, the sampling plane coincides with the sample contact surface.

[0010] In some embodiments, the sample contacting surface is made of black powder coated steel.

[0011] In some embodiments, the infrared imaging system further comprises one or more anesthesia ports configured for injecting anesthetic gas into the encapsulated housing and for collecting anesthetic gas from the encapsulated housing.

[0012] In some embodiments, the infrared imaging system further comprises a heating element in thermal contact with the sample holder.

[0013] In some embodiments, the infrared imaging system further comprises a barrier mounted to the sample holder, the barrier protruding upward from the sampling plane.

[0014] In some embodiments, each of the first lighting module and the second lighting module includes one or more laser diodes.

[0015] In some embodiments, the first infrared illumination beam and the second infrared illumination beam have a wavelength of about 750 nm, about 808 nm, or about 980 nm.

[0016] In some embodiments, the illumination area has a power of about 1 mW / mm 2 to about 3 mW / mm 2 Lighting power density within the range.

[0017] In some embodiments, each of the first lighting module and the second lighting module includes a Köhler integrator.

[0018] In some embodiments, the first lighting module and the second lighting module are symmetrically disposed on both sides of the detector.

[0019] In some embodiments, the first illumination module and the second illumination module are calibrated based on calibration data, which maps multiple orientations of the first illumination module and the second illumination module to corresponding multiple illumination power densities of the first infrared illumination beam and the second infrared illumination beam, and to corresponding multiple positions of the sample holder within the packaging shell.

[0020] In some embodiments, the detector comprises an InGaAs camera.

[0021] In some embodiments, the detector comprises:

[0022] sensor;

[0023] a first optical path device configured to collect and collimate light emitted by the fluorescent marker; and

[0024] The second optical path device is configured to form an image of the sample on the sensor.

[0025] In some embodiments, the infrared imaging system further comprises a motorized focus mechanism coupled to the detector, the motorized focus mechanism configured to vary a distance between the first optical path device and the second optical path device.

[0026] In some embodiments, the infrared imaging system further includes a filter wheel positioned between the first optical path device and the second optical path device, the filter wheel including a plurality of filters.

[0027] According to another aspect, a method for imaging a sample having a fluorescent marker is provided. The method includes: providing the sample on a sample holder having a sample contact surface and a sampling plane; generating a first infrared illumination beam and a second infrared illumination beam directed toward the sample using a first illumination module and a second illumination module, wherein the first infrared illumination beam and the second infrared illumination beam interact with each other at an imaging plane to form an illumination area having a rectangular shape and a uniform power distribution; moving the sample holder to a plurality of positions within a package; orienting the first infrared illumination beam and the second infrared illumination beam to move the illumination area within the package; causing the sampling plane and the imaging plane to overlap with each other at any of the plurality of positions within the package; and, when the sampling plane and the imaging plane overlap with each other, collecting light emitted by the fluorescent marker of the sample when the sample is illuminated by light from the illumination beam in the imaging plane.

[0028] In some embodiments, the method further comprises vertically offsetting the sampling plane from the sample contact surface.

[0029] In some embodiments, the sampling plane is vertically offset from the sample contact surface by a distance value corresponding to the thickness of the sample or the thickness of a portion of the sample.

[0030] In some embodiments, the sampling plane coincides with the sample contact surface.

[0031] In some embodiments, the method further comprises heating the sample holder.

[0032] In some embodiments, the first infrared illumination beam and the second infrared illumination beam have a wavelength of about 750 nm, about 808 nm, or about 980 nm.

[0033] In some embodiments, the method further includes modulating each of the first infrared illumination beam and the second infrared illumination beam using a Köhler integrator.

[0034] In some embodiments, the method further includes calibrating the first lighting module and the second lighting module based on calibration data, wherein the calibration data maps multiple orientations of the first lighting module and the second lighting module to corresponding multiple illumination power densities of the first infrared light beam and the second infrared light beam, and to corresponding multiple positions of the sample holder in the packaging shell.

[0035] In some embodiments, the method further comprises:

[0036] collecting and collimating light emitted by the fluorescent marker using a first optical path device; and

[0037] An image of the sample is formed on the sensor using the second optical path device.

[0038] Other features and advantages of the methods and systems described herein will be more readily understood when the preferred embodiments of the present invention are read with reference to the accompanying drawings. Although specific features described in the above summary of the invention and the following detailed description may be described with reference to specific embodiments or aspects thereof, it should be noted that these specific features may be combined with each other unless otherwise indicated. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 An embodiment of an infrared imaging system for imaging a sample having a fluorescent label is shown.

[0040] Figure 2 An embodiment of a lighting module is shown.

[0041] Figure 2A yes Figure 2 Cross-sectional view of the folding mirror 55 in FIG.

[0042] Figure 3 A schematic diagram of a detector according to one embodiment is shown.

[0043] Figure 4An example is shown where the sample holder begins in an initial or first position (left portion of this figure) and then moves to a subsequent or second position (right portion of this figure).

[0044] Figure 5 is a block diagram illustrating operable connections between some of the components included in an infrared imaging system. DETAILED DESCRIPTION

[0045] In this specification, similar features in the accompanying drawings have been given similar figure marks. In order to avoid confusing some of the drawings, if some elements have been identified in the previous drawings, they may no longer be indicated. It should be understood that the elements of the drawings are not necessarily depicted to scale, because the emphasis is placed on clearly showing the elements and structures of the embodiments of the present application. In addition, for easy and clear description, position descriptors indicating the position and / or direction of an element relative to another element are used herein. Unless otherwise indicated, these position descriptors should be used with reference to the context of the accompanying drawings and should not be considered to be restrictive. More specifically, it should be understood that, in addition to the orientations illustrated in the drawings, such spatially relative terms are intended to cover various different orientations during use or operation of the embodiments of the present application.

[0046] Unless otherwise indicated, the terms "connected" and "coupled," and their derivatives and variations, as used herein, refer to a direct or indirect structural or functional connection or coupling between two or more elements. For example, the connection or coupling between elements can be mechanical, optical, electrical, thermal, logical, or any combination thereof.

[0047] Unless otherwise indicated, the terms "a," "an," and "an" are defined herein as "at least one," ie, these terms do not exclude plural referents.

[0048] Terms such as "substantially", "typically" and "about" used to modify the values, conditions or characteristics of the features of the exemplary embodiments should be understood as meaning that the values, conditions or characteristics are defined within an acceptable tolerance range for the correct operation of the exemplary embodiment with respect to its intended application, or fall within an acceptable range of experimental error. Specifically, the term "approximately" generally refers to a numerical range that a person skilled in the art would consider to be equivalent to the value (e.g., having the same or equivalent function or effect). In some cases, the term "approximately" means a variation of ±10% of the value described. It should be noted that, unless otherwise indicated, all numerical values ​​used herein are assumed to be modified by the term "approximately".

[0049] Likewise, the terms "superimposition," "superimpose," "superimposed," and "superimposing" are intended herein to mean that two elements are either in the same position or within a certain predetermined tolerance range with respect to spatial alignment with each other. That is, these terms are intended to encompass not only overlapping two elements "exactly" or "identically," but also overlapping two elements "substantially," "approximately," or "subjectively," as well as providing a higher or best overlap among multiple overlap possibilities.

[0050] In this specification, the expression "based on" is intended to mean "based at least in part on," that is, the expression may mean "based only on" or "based in part on," and should not be interpreted in a limiting manner. More specifically, the expression "based on" may also be understood to mean "depending on," "indicative of," "associated with," or similar expressions.

[0051] In this specification, the terms "light" and "optical" and their variations and derivatives are used to refer to radiation in any suitable segment of the electromagnetic spectrum. The terms "light" and "optical" are therefore not limited to visible light, and may also include, without limitation, infrared and ultraviolet segments. For example, in some embodiments, the technology of the present application may be used with an electromagnetic signal having a wavelength of about 400 nm to about 1700 nm, for example, between 1000 nm and 1700 nm. However, this range is provided for illustrative purposes only, and some embodiments of the technology of the present application may operate outside this range. At the same time, the skilled person will be aware that the specific definitions of the ultraviolet, visible, infrared, and near-infrared ranges in terms of spectral ranges and the dividing lines therebetween may vary depending on the technical field or definition under consideration, and are not intended to limit the scope of application of the technology of the present application.

[0052] In this specification, the expression "illumination beam spectrum" and its synonyms or derivatives are used in a broad sense to refer to the spectral power distribution of the illumination beam. The illumination spectrum can be expressed as the radiant power distribution per unit area and per unit wavelength or frequency over a spectral region of the electromagnetic spectrum.

[0053] This specification generally relates to infrared imaging systems and one or more associated methods and techniques for preclinical imaging purposes. In the context of this specification, infrared imaging systems may sometimes be referred to as "IR VIVO instruments." It is important to note that the term "preclinical imaging" is understood herein to refer to techniques that allow for visualization and examination of live animals (e.g., small animals such as mice and rats). Preclinical imaging techniques may be particularly useful for research purposes (e.g., drug development).

[0054] The infrared imaging system described in more detail below is a fluorescence-based imaging instrument. Such instruments typically include a light source that uses excitation light to excite fluorescent probes within a sample, which in the context of a preclinical imaging system may be a small animal. Such instruments also include a detector configured to detect the fluorescent signals generated by these probes. Other optical components may be provided between the sample and the detector, such as, for example and without limitation, one or more imaging lenses, one or more spectral filters, a dichroic element (e.g., for separating the output signal into two spectral bands that can be detected by two different cameras), and other optical components.

[0055] Turning now to the figures, various embodiments of infrared imaging systems and methods will be presented.

[0056] refer to Figure 1 , which shows an infrared imaging system 20 for imaging a sample 22. It is noteworthy that the sample 22 may include one or more animals. Likewise, the expression "sample" is not limited to referring to only one animal and is not intended to be limiting. A fluorescent marker (not shown) is provided in the sample 22. For example, and without limitation, the fluorescent marker may be injected into the small animal being imaged. It is noteworthy that the expressions "fluorescent marker" and "fluorescent probe" will be used interchangeably throughout this specification. Non-limiting examples of fluorescent markers are quantum dots (e.g., PbS, Ag2S), organic molecules such as indocyanine green (ICG) and IR800 dye molecules, single-walled carbon nanotubes, rare earth nanoparticles, and the like.

[0057] Package housing and sample holder

[0058] Infrared imaging system 20 includes an enclosure 24 (sometimes referred to as a "chamber"). Enclosure 24 includes walls defining an interior volume 26 within which at least some of the other components of infrared imaging system 20 may be mounted. Enclosure 24 typically includes a door or drawer (not shown) for accessing interior volume 26 (and its contents) when desired. Figure 1 ), for example, and without limitation, for creating preclinical tests and preparing samples 22. In some embodiments, the entire enclosure 24 can be light-tight. In some embodiments, the doors or drawers are light-tight. Notably, the enclosure 24 can be equipped with components, such as, for example and without limitation, one or more anesthetic gas manifolds, one or more gas conduits, and one or more hot plates, as will be described in more detail below.

[0059] The infrared imaging system 20 also includes a sample holder 28. The sample holder 28 is positioned in the encapsulating housing 24 and has a sample contact surface 30. The sample contact surface 30 has a sampling plane 32. Notably, in some embodiments, the sampling plane 32 may be vertically offset from the sample contact surface 30 by a distance value corresponding to the thickness (or height) of the imaged animal or a portion of the thickness (or height) of the imaged animal. Alternatively, the sampling plane 32 may coincide with or substantially coincide with the sample contact surface 30. The sample 22 may be placed on the sample contact surface 30 so that the sampling plane 32 intersects the sample 22 or at least a portion of the sample 22. As will be described in more detail below, the sample holder 28 can be translated and adjusted along three dimensions, such as the x-axis, the y-axis, and the z-axis. Similarly, the position of the sampling plane 32 can be changed or adjusted.

[0060] In some embodiments, sample contact surface 30 is made of a steel plate coated with black powder. In other embodiments, sample contact surface 30 may be made of aluminum. Alternatively, sample contact surface 30 may be made of any type of material having suitable properties for preclinical testing and related medical applications (e.g., anodized aluminum, heated glass, etc.), as long as the material has low reflectivity and low fluorescence in the infrared band and can be relatively easily cleaned using an ethanol solution or bleach. Notably, the material forming sample contact surface 30 can be selected based on chemical properties (e.g., composition) and / or physical properties (e.g., optical and magnetic properties).

[0061] The infrared imaging system 20 generally includes components found in typical preclinical equipment. For example, the infrared imaging system 20 may generally include one or more anesthesia ports. In some embodiments, the infrared imaging system 20 includes three anesthesia ports, meaning that three samples 22 can be placed on the sample contact surface 30. It should be noted that the number of anesthesia ports may vary from three, and the fact that the infrared imaging system 20 may include three anesthesia ports is for illustrative purposes only and should not be considered limiting. For example, the infrared imaging system 20 may include one, two, three, four, five, or more anesthesia ports. Similarly, the number of animals comprising the sample 22 may vary from three. For example, and without limitation, the sample 22 may include one, two, three, four, five, or more animals. In some embodiments, the number of anesthesia ports may be equal to the number of animals comprising the sample 22. In other embodiments, the number of anesthesia ports may be proportional to, or at least correlated with, the number of animals comprising the sample 22. For example, and without limitation, the ratio of the number of anesthesia ports to the number of animals comprising the sample 22 may be 1:1, 1:2, 1:3, or any other ratio that allows the anesthesia ports to fulfill their function. In this regard, it is noteworthy that the anesthesia port allows for the injection and collection of anesthetic gas for input and output of anesthetic gas from the enclosure 24. Anesthetic gas is often used to hold the sample 22 stationary during imaging.

[0062] Infrared imaging system 20 may also include a hot plate or similar device. Notably, the hot plate can maintain the entire interior volume 26 or only a portion thereof (e.g., sample contact surface 30) at a given temperature. The hot plate can generally be used to maintain sample 22 at a given temperature. In fact, in the case of small animals or mammals in general, their average body temperature tends to drop while under anesthesia. The hot plate can thus mitigate this effect. Infrared imaging system 20 may also include a barrier or fence mounted to or on sample holder 28. This feature can be particularly useful in the rare event that sample 22 (e.g., a small animal) awakens while under anesthesia or in the event of a failure during anesthesia.

[0063] The size and geometry of sample holder 28 may vary. However, the size of sample holder 28 is preferably such that sample holder 28 generally fits within the field of view of imaging system 20, as will be described in more detail below. In one example of an embodiment, the field of view has the following dimensions: approximately 15.6 cm x approximately 12.5 cm.

[0064] In some embodiments, the sample holder 28 may include a lower platform and an upper platform. The lower platform may span the entire floor (i.e., width and depth) of the packaging housing 24. The upper platform may be smaller and may have the following dimensions: 300 mm × 250 mm. The upper platform may be mounted to the lower platform. In some embodiments, the upper platform may be mechanically connected to the lower platform via a two-dimensional translation stage. In the context of the present description, the two-dimensional translation stage is configured to translate the surface of the upper platform along the X-axis and the Y-axis to move the sample 22 to one side, along these two axes, and relative to the field of view of the camera.

[0065] light source

[0066] The infrared imaging system 20 includes a light source 34 configured to illuminate the sample contact surface 30. The light source 34 includes a first lighting module 36 and a second lighting module 38. The first lighting module 36 and the second lighting module 38 are each configured to project a corresponding first infrared illumination beam 40 and a second infrared illumination beam 42 toward the sample holder 30. The first lighting module 36 and the second lighting module 38 can each generate a relatively high power infrared illumination beam. It is important to note that the first lighting module 36 and the second lighting module 38 can each include one or more laser diodes, each laser diode being associated with a corresponding optical characteristic (e.g., intensity and / or spectral distribution). The relatively high power can be used to generate a fluorescent signal that is strong enough to be detected (approximately 0.05 mW / mm 2 to about 3 mW / mm 2 ). It should be noted that fluorescent markers used in the NIR-II portion of the electromagnetic spectrum generally have relatively low efficiency compared to fluorescent markers used in the visible portion of the electromagnetic spectrum. It should be noted that detectors configured to operate in the NIR-II portion of the electromagnetic spectrum are generally less sensitive than detectors configured to operate in the visible portion of the electromagnetic spectrum. It should be noted that the wavelengths of the first infrared illumination beam 40 and the second infrared illumination beam 42 emitted by the first illumination module 36 and the second illumination module 38 can be selected and varied. The selection can be manual (e.g., by a user) or automatic (e.g., each module 36, 38 can select an infrared illumination wavelength sequentially and / or automatically). This feature is useful for exciting fluorescent markers of different properties. It will be noted that illumination of different wavelengths is generally performed sequentially, rather than simultaneously. Non-limiting examples of wavelengths that can be used are 750 nm, 808 nm, 860 nm, and 980 nm.

[0067] First infrared illumination beam 40 and second infrared illumination beam 42 interact at imaging plane 44 to form illumination area 46. Imaging plane 44 extends along the X-axis and the Y-axis. Illumination area 46 has a rectangular shape and a uniform power distribution (sometimes referred to as "illumination distribution" or "power density distribution"), and also extends along the X-axis and the Y-axis. The power or illumination density in the illumination area represents the cumulative power of first infrared illumination beam 40 and second infrared illumination beam 42. In some embodiments, for a total power of about 20 W, the power or illumination density is about 1 mW / mm 2 These values ​​are relatively close to but lower than the limit of power or illumination density for living tissue (about 3mW / mm 2 ). Those skilled in the art will note that illumination in existing preclinical imagers that use visible light tubes is typically provided by halogen white lamps. These lamps are typically optically coupled to filters (e.g., filter wheels) for filtering the excitation light. Existing preclinical imagers that use visible light may also use LED light sources and / or laser light sources. In these cases, the illumination density of the existing imagers is less than that which can be achieved using the first illumination module 36 and the second illumination module 38 included in the technology described herein. In fact, the illumination density that can be achieved using the technology presented in the disclosure of this application is an order of magnitude higher than the illumination density that can be achieved using existing technologies that rely on halogen devices and LED devices.

[0068] The light generated by each laser diode included in the first lighting module 36 and the second lighting module 38 is optically structured via the optical components included in the first lighting module 36 and the second lighting module 38. This allows an illumination area 46 with a substantially uniform rectangular shape to be formed on the sample 22. It will be noted that the illumination area 46 is generally of the same size as the field of view of the infrared imaging system 20. Those skilled in the art will appreciate that uniform illumination is important to ensure that each part of the sample 22 receives the same illumination density. For example, in a case where three mice are imaged and placed in the field of view, they must be illuminated using the same power density so that the imaging results can be compared with each other. It is also advantageous to limit the illumination area 46 to the field of view of the infrared imaging system 20 to avoid wasting laser power. In fact, if too much laser power is lost, more high-power lasers will be needed, which will increase the cost of the system and also increase the complexity of the thermal management of the first lighting module 36 and the second lighting module 38. Furthermore, those skilled in the art will note that projecting only a portion of first infrared illumination beam 40 and second infrared illumination beam 42 will result in an increase in unnecessary stray light detectable by the detector in infrared imaging system 20 and will thereby reduce the overall sensitivity of infrared imaging system 20.

[0069] Now turn Figure 2 The first lighting module 36 and the second lighting module 38 use the design of a Köhler integrator to structure the light emitted by the laser diode. Figure 2 In the figure, only the first illumination module 36 is shown, and it will be readily understood that the description also applies to the second illumination module 38. The design of a Köhler integrator is known in the art and uses elements including a collimating lens 48, a first fly's eye lens 50, a second fly's eye lens 52, a projection lens 54, and a folding mirror 55. These elements define an optical path therebetween, which extends along an optical axis 56. In some embodiments, the projection lens 54 may be replaced by other optical components (not shown). Such other optical components may include, but are not limited to, lenses, mirrors, filters, and other suitable reflective, refractive, and / or diffractive optical components.

[0070] In the illustrated embodiment, first illumination module 36 and second illumination module 38 are positioned to project light (ie, first infrared illumination beam 40 and second infrared illumination beam 42) from above sample 22. Referring now to Figure 1 , it can be seen that the first lighting module 36 and the second lighting module 38 are located on either side of the detector 58. In some embodiments, the first lighting module 36 and the second lighting module 38 are symmetrically disposed on either side of the detector 58. However, it will be understood that the first lighting module 36 and the second lighting module 38 may be disposed at other locations relative to the detector and / or may be asymmetrically arranged relative to the detector without departing from the scope of what is claimed.

[0071] First lighting module 36 and second lighting module 38, and therefore first infrared illumination beam 40 and second infrared illumination beam 42, are not substantially parallel to the Z-axis, but form a small angle with respect to the Z-axis. When sample 22 includes multiple animals (i.e., two or more animals), it is important to note that in order to prevent the animals from casting shadows on each other, the angles between the first illumination beam and the second illumination beam and the Z-axis are kept as small as possible.

[0072] As will be described in more detail below, first illumination module 36 and second illumination module 38 are rotatable, which allows, for example, first infrared illumination beam 40 and second infrared illumination beam 42 to be projected at or near the center of the field of view of infrared imaging system 20 as sample holder 28 moves upward and / or downward. Rotation of first illumination module 36 and second illumination module 38 allows uniform illumination in illumination area 46 to be maintained.

[0073] Motor components

[0074] Figure 5A block diagram illustrating operable connections between some of the components included in infrared imaging system 20. Infrared imaging system 20 includes a motor assembly 60 configured to move a sample holder to a plurality of positions within enclosure 24. Motor assembly 60 may include one or more motors. The motors may be of any type or design.

[0075] In some embodiments, the sample holder 28 can be moved or translated along two axes (e.g., an X-axis and a Y-axis) by the motor assembly 60. The motor assembly 60 can be configured to translate the sample holder 28 sequentially or simultaneously in the X-axis and Y-axis directions. For example, the sample holder 28 can be sequentially translated first in a direction parallel to the X-axis and then in a direction parallel to the Y-axis, or vice versa. Alternatively, the sample holder 28 can be configured to be simultaneously adjustable along the X-axis and the Y-axis. It will be noted that the motor assembly 60 can monitor or record the displacement of the sample holder 28 along each axis. The monitored or recorded information is included in the calibration data.

[0076] In some embodiments, the motor assembly 60 can include two motors, each of which is configured to move the sample holder 28 along a corresponding direction (eg, the X-axis or the Y-axis).

[0077] The sample holder 28 can also be moved or translated in the Z-axis direction. This displacement of the sample holder 28 in the Z-axis direction generally occurs once the sample holder 28 has been aligned or positioned in the X-axis and Y-axis directions within the enclosure 24. Movement of the sample holder 28 in this direction can be provided by one or more motors.

[0078] It will be appreciated that movement of the sample holder 28 using the motor assembly 60 allows for transitioning from a "wide field of view mode" to a "close field of view mode." Switching between these two field of view modes may be useful when the sample 22 includes multiple animals. For example, and without limitation, the motor assembly 60 may allow for transitioning from a first field of view encompassing all of the animals in the sample 22 to a second field of view encompassing only one of the animals forming the sample 22 or forming a portion of the sample 22. In some embodiments, the field of view may be adjusted to simultaneously image a portion of each of the animals forming the sample 22, which may be useful in situations where characteristics of specific portions of the various animals are to be compared.

[0079] In some embodiments, the motor assembly 60 is manually operated. In these embodiments, translating the sample holder 28 involves two steps. In the first step, a spring-loaded, normally-open brake that prevents the upper platform from sliding is released. This brake can be temporarily released by pressing a button, or semi-permanently released by pressing and locking the same button. Once the brake is released, the upper platform can be moved relatively smoothly "side to side" (e.g., along the X-axis) and "forward and backward" (e.g., along the Y-axis). In other embodiments, the motor assembly 60 is automated.

[0080] Optomechanical mechanism

[0081] refer to Figure 5 Infrared imaging system 20 includes an optomechanical mechanism 62 configured to adjust the orientation of first and second infrared illumination beams 40, 42 to move illumination area 46 within enclosure 24. Adjusting the orientation of first and second infrared illumination beams 40, 42 generally includes changing the spatial configuration of first and second illumination modules 36, 38 (via rotation, translation, or a combination thereof). More specifically, adjustment of the orientation of first and second infrared illumination beams 40, 42 by optomechanical mechanism 62 in conjunction with motor assembly 60 may be used to control the size of illumination area 46.

[0082] In some embodiments, and with reference now to Figure 2 , adjusting the orientation of first illumination module 36 and second illumination module 38 includes rotating first illumination module 36 and second illumination module 38 about optical axis 56. Rotating first illumination module 36 and second illumination module 38 from the initial position toward the subsequent position causes the interaction plane of first infrared illumination beam 40 and second infrared illumination beam 42 to move from the first imaging plane (associated with the initial position of modules 36, 38) to the subsequent imaging plane (associated with the subsequent position of modules 36, 38), thereby forming a subsequent illumination area that also has a generally rectangular shape and a uniform power distribution, as shown in reference to FIG. Figure 4 Will be described in more detail. It is noteworthy that the first imaging plane and the subsequent imaging plane are generally not located at the same position along the Z axis. For example, the subsequent imaging plane is generally higher or lower than the first imaging plane. Similarly, the orientation adjustment of the first lighting module 36 and the second lighting module 38 results in a change in the position of the imaging plane 44 within the package shell 24 (for example, along the Z axis). In other embodiments, only the folding reflector 55 can be rotated. It is noteworthy that in addition to adjusting the orientation thereof, the power of the first infrared illumination beam 40 and the second infrared illumination beam 42 can also be changed (i.e., modified or changed). The operation of changing or adjusting the orientation of the first infrared illumination beam 40 and the second infrared illumination beam 42 to move the illumination area 46 within the package shell is generally referred to as "illumination modulation."

[0083] It must be noted that the first lighting module 36 and the second lighting module 38 are generally calibrated. The calibration data includes, but is not limited to, a mapping between a plurality of orientations of the first lighting module 36 and the second lighting module 38 and the corresponding optical properties of the first infrared illumination beam 40 and the second infrared illumination beam 42. Similarly, once the position of the sample holder 28 in the packaging shell 24 is known, the orientation of the first lighting module 36 and the second lighting module 38 to be achieved can be determined because this mapping information is contained in the calibration data or can be calculated based on the calibration data, for example, by interpolation, extrapolation, and other methods. The interpolation method can be linear, polynomial (Lagrange, Newton, etc.), curve fitting, etc. In one example, the calibration step is used to maintain approximately 1 mW / mm at any position within the packaging shell 24. 2 to about 3mW / mm 2 A relatively constant power density is maintained even when the distance between sample holder 28 and detector 58 changes. More specifically, changes in the orientation of first infrared illumination beam 40 and second infrared illumination beam 42 allow a rectangular and uniform power distribution to be maintained at least partially, approximately, or substantially at any location within enclosure 24. In some embodiments, the calibration data may also include information regarding the illumination power that needs to be provided or generated in order to maintain a constant or desired power density. Note that the power density generally increases as sample holder 28 is raised, i.e., as the distance between sample holder 28 and first illumination module 36 and second illumination module 38 decreases. The rectangular and uniform illumination area becomes smaller as sample holder 28 is raised. In some embodiments, the power density of first infrared illumination beam 40 and second infrared illumination beam 42 can be controlled to maintain a relatively constant power density across the illumination area. In some embodiments, the power density of first illumination module 36 and second illumination module 38 can be constant as the platform is raised, which can increase the illumination power density across the illumination area.

[0084] control unit

[0085] As in Figure 5As shown in FIG, infrared imaging system 20 includes a control unit 64. Control unit 64 is operably connected to motor assembly 60 and optomechanical mechanism 62. Control unit 64 is configured to cause sampling plane 32 and imaging plane 44 to overlap at any one of a plurality of positions within enclosure 24. When sampling plane 32 and imaging plane 44 overlap, the two planes are located at substantially the same position along the Z axis. Notably, when sampling plane 32 is vertically offset from sample contact surface 30, such as when a distance is required to accommodate the thickness of sample 22 (or a portion thereof), control unit 64 can be configured to position sampling plane 32 and imaging plane 44 based on this vertical offset. In some embodiments, the value of the vertical offset can be automatically determined by infrared imaging system 20. In other embodiments, the value of the vertical offset can be provided by a user, such as manually. In yet another embodiment, the value of the vertical offset can be obtained from a database. Such a database can, for example and without limitation, associate average values ​​of the thickness of one or more animals forming sample 22 with corresponding positions of sample holder 28 within enclosure 24.

[0086] For example, and without limitation, the control unit 64 can be implemented by a programmable computer comprising at least one processor, a data storage system (including volatile and non-volatile memory and / or storage elements), at least one input device, and at least one output device. In some embodiments, the programmable computer can execute a computer program that allows it to control the motor assembly 60 and the optomechanical mechanism 62. The control unit 64 is configured to track, monitor, or record the position of the sample holder 28 within the encapsulating housing 24. More specifically, the control unit 64 receives the position of the sample holder 28 within the encapsulating housing 24 as input and, based on calibration data, outputs a signal for transmission to the first and second illumination modules 36 and 38.

[0087] Reference Figure 4 , shows an example in which the sample holder 28 begins in an initial or first position (left portion of this figure) and then moves to a subsequent or second position (right portion of this figure).

[0088] In the initial position, sample holder 28 is positioned so that the distance between detector 58 and sample holder 28 is approximately 400 mm (in the Z-axis direction). Similarly, sampling plane 32 is approximately 400 mm away from detector 58. In the initial position, the orientations of first illumination module 36 and second illumination module 38 are adjusted so that illumination area 46 covers sample 22 and imaging plane 44 and sampling plane 32 overlap.

[0089] In the subsequent position, the sample holder 28 is positioned so that the distance between the detector 58 and the sample holder 28 is about 200 mm (in the Z-axis direction), which means that the sample holder 28 has been brought closer to the detector 58. Likewise, the sampling plane 32' is about 200 mm away from the detector 58. In the subsequent position, the orientation of the first lighting module 36 and the second lighting module 38 is changed compared to the initial position. It will be noted that in Figure 4 In the subsequent position shown in the non-limiting embodiment of FIG, the orientations of first illumination module 36 and second illumination module 38 are adjusted so that illumination area 46 ′ covers sample 22 and imaging plane 44 ′ and sampling plane 32 overlap each other.

[0090] It should be noted that the shape and size of the illumination area 46 may also change when the orientation of the first illumination module 36 and the second illumination module 38 are changed and / or the position of the sample holder 28 within the packaging housing 24 is changed.

[0091] detector

[0092] As mentioned above, infrared imaging system 20 includes detector 58, see for example Figure 1 and Figure 3 Detector 58 is configured to receive light emitted by fluorescent markers of sample 22 when sample 22 is illuminated in imaging plane 44 by first infrared illumination beam 40 and second infrared illumination beam 42 when sampling plane 32 and imaging plane 44 overlap. In the depicted embodiment, detector 58 comprises an InGaAs camera. Such a camera allows for relatively precise localization of fluorescence originating from within sample 22, providing valuable information to biologists.

[0093] As in Figure 3 As better shown in FIG, the detector 58 includes two optical paths 66, 68 separated by a filter wheel 70. The detector 58 also includes a sensor 72.

[0094] The first optical path device 66 collects light emitted from the sample 22 and roughly collimates the light. The resulting light then passes through a dual filter wheel 70. The filter wheel 70 generally includes a plurality of filters. In some embodiments, the filter wheel 70 includes a bandpass filter and an edgepass filter. In some embodiments, the first optical path device 66 may include one or more detection lenses and / or any other optical elements. For example, in one embodiment, the first optical path device 66 may include two lenses. The second optical path device 68 forms an image of the sample 22 on the sensor 72. In some embodiments, the second optical path device 68 may include one or more lenses and / or any other optical elements. In some embodiments, the detector 58 is provided with a motorized focusing mechanism 74. The motorized focusing mechanism 74 is positioned between the second optical path device 68 and the sensor 72 and is capable of changing the distance between the second optical path device 68 and the sensor 72 to adjust the focal length.

[0095] The filters included in filter wheel 70 are typically dielectric interference filters whose transmission wavelength is angle-dependent. In some embodiments, the optical design of first light path device 66 is such that light passing through filter wheel 70 is transmitted as close to normal incidence as possible (i.e., parallel to the Z-axis). Light from different viewpoints on sample 22 strikes the filters at different angles. Similarly, optimizing the optical design of first light path device 66 to allow light to pass through the filters at close to normal incidence ensures that the wavelengths detected for each point in the field of view are very close to each other.

[0096] Note also that positioning the filter wheel 70 between the two optical path devices 66 , 68 also prevents stray light from striking the filters at angles away from normal, which would result in unwanted light not being blocked by the filters and thereby producing undesirable artifacts in the image formed on the sensor 72 .

[0097] In conjunction with the operation of motor assembly 60 and optomechanical mechanism 62, varying the working distance (i.e., the distance between sample 22 and sensor 72) and adjusting the focus of infrared imaging system 20 (e.g., using the motorized focus mechanism) allows imaging of various areas, ranging from an area measuring approximately 156 mm by approximately 125 mm (i.e., illumination area 46) to an area measuring approximately 50 mm by approximately 40 mm (i.e., illumination area 46). This means that the field of view of detector 58 can be controlled. In the first configuration, up to three mice (or other similar samples 22) can be imaged. In the second configuration, approximately one-third of the body of a single mouse (or similar sample 22) can be imaged within an area measuring 50 mm by 40 mm (at a Z-axis distance of 200 mm).

[0098] The combination of illumination modulation, XY translation, and field of view (FOV) adjustment allows the infrared imaging system to capture images of the sample 22 at any position in a three-dimensional space of 156 mm × 125 mm × 50 mm, with a spatial sampling rate of 80 μm per pixel when the field of view is approximately 50 mm × 40 mm.

[0099] method

[0100] According to an embodiment, a method of imaging a sample having a fluorescent label is also provided.

[0101] The method may include the step of providing a sample on a sample holder having a sample contact surface and a sampling plane.

[0102] The method may include the steps of generating a first infrared illumination beam and a second infrared illumination beam directed toward the sample using a first illumination module and a second illumination module, wherein the first infrared illumination beam and the second infrared illumination beam interact at an imaging plane to form an illumination area having a rectangular shape and uniform power distribution.

[0103] The method may include the step of moving the sample holder to a plurality of positions within the enclosure.

[0104] The method may include the step of adjusting the orientation of the first infrared illumination beam and the second infrared illumination beam to move the illumination area within the enclosure.

[0105] The method may include the step of causing the sampling plane and the imaging plane to overlap with each other at any one of a plurality of locations within the encapsulating housing.

[0106] The method may comprise the step of collecting light emitted by fluorescent markers of the sample when the sample is illuminated by light of the illumination beam in the imaging plane when the sampling plane and the imaging plane overlap each other.

[0107] In some embodiments, the method further comprises vertically offsetting the sampling plane from the sample contact surface.

[0108] In some embodiments, the sampling plane is vertically offset from the sample contact surface by a distance value corresponding to the thickness of the sample or a thickness of a portion of the sample.

[0109] In some embodiments, the sampling plane coincides with the sample contact surface.

[0110] In some embodiments, the method further comprises heating the sample holder.

[0111] In some embodiments, the first infrared illumination beam and the second infrared illumination beam have a wavelength of about 750 nm, about 808 nm, or about 980 nm.

[0112] In some embodiments, the method further includes modulating each of the first infrared illumination beam and the second infrared illumination beam using a Köhler integrator.

[0113] In some embodiments, the method further includes calibrating the first lighting module and the second lighting module based on calibration data, wherein the calibration data maps multiple orientations of the first lighting module and the second lighting module to corresponding multiple illumination power densities of the first infrared light beam and the second infrared light beam, and maps to corresponding multiple positions of the sample holder in the packaging shell.

[0114] In some embodiments, the method further comprises collecting and collimating light emitted by the fluorescent marker using a first optical path device and forming an image of the sample on a sensor using a second optical path device.

[0115] Some alternative embodiments and examples have been described and illustrated herein. The embodiments described above are intended to be exemplary only. Those skilled in the art will appreciate the features of each individual embodiment and the possible combinations and variations of the components. Those skilled in the art will also understand that any embodiment can be combined in any way with other embodiments disclosed herein. Therefore, the examples and embodiments of the present application can be considered in all respects to be illustrative and not restrictive. Therefore, although specific embodiments have been shown and described, a variety of variant embodiments can be envisioned without significantly departing from the scope defined in the appended claims.

Claims

1. An infrared imaging system for imaging a sample having a fluorescent label, the infrared imaging system comprising: Encapsulation shell; A sample holder is installed in the packaging shell, and the sample holder has a sample contact surface and a sampling plane; a light source configured to illuminate the sample contact surface, the light source comprising a first illumination module and a second illumination module, the first illumination module and the second illumination module each being configured to project a corresponding first infrared illumination beam and a second infrared illumination beam toward the sample holder, the first infrared illumination beam and the second infrared illumination beam interacting at an imaging plane to form an illumination area having a rectangular shape and a uniform power distribution, wherein the first illumination module and the second illumination module are calibrated based on calibration data, the calibration data mapping a plurality of orientations of the first illumination module and the second illumination module to a corresponding plurality of illumination power densities of the first infrared illumination beam and the second infrared illumination beam, and to a corresponding plurality of positions of the sample holder within the enclosure; a motor assembly configured to move the sample holder to a plurality of positions within the enclosure; an optomechanical mechanism configured to adjust the orientation of the first infrared illumination beam and the second infrared illumination beam to move the illumination area within the package housing; a control unit operatively connected to the motor assembly and the optomechanical mechanism, the control unit being configured to cause the sampling plane and the imaging plane to overlap with each other at any one of the plurality of positions within the enclosure; and a detector configured to receive light emitted by the fluorescent markers of the sample when the sample is illuminated in the imaging plane when the sampling plane and the imaging plane overlap each other, wherein the calibration data ensures a rectangular shape and uniform power distribution of the illumination area even if a distance between the sample holder and the detector varies.

2. The infrared imaging system of claim 1, wherein the encapsulating housing defines an interior volume, the encapsulating housing further comprising a door or drawer for accessing contents of the interior volume.

3. An infrared imaging system according to claim 1 or 2, wherein the sampling plane is vertically offset from the sample contact surface.

4. The infrared imaging system of claim 3, wherein the sampling plane is vertically offset from the sample contact surface by a distance value corresponding to the thickness of the sample or a thickness of a portion of the sample.

5. The infrared imaging system according to claim 1 or 2, wherein the sampling plane coincides with the sample contact surface.

6. The infrared imaging system of claim 1, wherein the sample contact surface is made of black powder-coated steel. 7 . The infrared imaging system of claim 1 , further comprising one or more anesthesia ports configured for injecting anesthesia gas into the enclosure and for collecting the anesthesia gas from the enclosure.

8. The infrared imaging system of claim 1 further comprising a heating element in thermal contact with the sample holder.

9. The infrared imaging system of claim 1, further comprising a barrier mounted to the sample holder, the barrier protruding upward from the sampling plane.

10. The infrared imaging system of claim 1 wherein each of the first and second illumination modules comprises one or more laser diodes.

11. The infrared imaging system of claim 1 wherein the first infrared illumination beam and the second infrared illumination beam have wavelengths of approximately 750 nm, approximately 808 nm, or approximately 980 nm.

12. The infrared imaging system of claim 1 , wherein the illumination area has a power of from about 1 mW / mm 2 to about 3mW / mm 2 lighting power density within the range of .

13. The infrared imaging system of claim 1, wherein each of the first illumination module and the second illumination module comprises a Köhler integrator. 14 . The infrared imaging system according to claim 1 , wherein the first lighting module and the second lighting module are symmetrically arranged on both sides of the detector.

15. The infrared imaging system of claim 1 wherein the detector comprises an InGaAs camera.

16. The infrared imaging system of claim 1 , wherein the detector comprises: sensor; a first optical path device, configured to collect and collimate light emitted by the fluorescent marker; as well as The second optical path device is configured to form an image of the sample on the sensor.

17. The infrared imaging system of claim 16, further comprising a motorized focus mechanism coupled to the detector, the motorized focus mechanism configured to vary a distance between the first optical path device and the second optical path device.

18. The infrared imaging system of claim 16 or 17, further comprising a filter wheel positioned between the first optical path device and the second optical path device, the filter wheel comprising a plurality of filters.

19. A method for imaging a sample having a fluorescent label, the method comprising: providing a sample on a sample holder having a sample contact surface and a sampling plane; generating, using a first illumination module and a second illumination module, a first infrared illumination beam and a second infrared illumination beam directed toward the sample, wherein the first infrared illumination beam and the second infrared illumination beam interact with each other at an imaging plane to form an illumination area having a rectangular shape and uniform power distribution; calibrating the first illumination module and the second illumination module based on calibration data, wherein the calibration data maps a plurality of orientations of the first illumination module and the second illumination module to a corresponding plurality of illumination power densities of the first infrared beam and the second infrared beam, and to a corresponding plurality of positions of the sample holder within the packaging housing; moving the sample holder to a plurality of positions within the packaging housing; Adjusting the orientations of the first infrared illumination beam and the second infrared illumination beam to move the illumination area within the packaging housing; causing the sampling plane and the imaging plane to overlap with each other at any one of the plurality of positions within the packaging housing; as well as When the sampling plane and the imaging plane overlap each other, light emitted by the fluorescent markers of the sample when the sample is illuminated by light of the illumination beam in the imaging plane is collected, wherein the calibration data ensures a rectangular shape and a uniform power distribution of the illumination area even if the distance between the sample holder and the detector changes.

20. The method of claim 19, further comprising vertically offsetting the sampling plane from the sample contact surface.

21. The method of claim 20, wherein the sampling plane is vertically offset from the sample contact surface by a distance value corresponding to the thickness of the sample or a thickness of a portion of the sample.

22. The method of claim 19, wherein the sampling plane coincides with the sample contact surface.

23. The method of claim 19, further comprising heating the sample holder.

24. The method of claim 19, wherein the first infrared illumination beam and the second infrared illumination beam have a wavelength of approximately 750 nm, approximately 808 nm, or approximately 980 nm.

25. The method of claim 19, further comprising modulating each of the first and second infrared illumination beams using a Köhler integrator.

26. The method of claim 19, further comprising: collecting and collimating light emitted by the fluorescent marker using a first optical path device; An image of the sample is formed on a sensor using a second optical path device.

Images