Apparatus for thermostatically operating a live animal for internal tissue microscopic imaging

Abstract

The present invention relates to a device for immobilization and body temperature maintenance for immobilizing a small animal for facilitating generation of high resolution microscopic images of predetermined tissue of said small animal within a biological microscope having an objective, said device comprising: a plate heater having a heating wire mounted therein, enabling direct placement of said small animal on said plate heater; a glass heater holder fixed to a hole of said plate heater; and a glass heater fixed to said glass heater holder and located above said tissue of said small animal, and maintaining planarity of said tissue and increasing temperature of said tissue itself, wherein a cover glass for adjusting refractive index of said objective is integrally formed with the heating wire heater. In an embodiment of the present invention, the body temperature of a live small animal as an object to be observed with a biological microscope is maintained constant.

Description

TECHNICAL FIELD

[0001] This application claims priority to Korean Patent Application No. 10-2021-0107411 filed on August 13, 2021, the entire contents of which are incorporated herein by reference.

[0002] The present invention relates to bio-imaging of pre-clinical animal models, and in particular to a fixation and body temperature maintenance apparatus to which various auxiliary devices are easily attachable and detachable, and which fixes a live animal and maintains the body temperature of the live animal when imaging internal tissues of the live animal in a biological microscope or an endoscopic microscope. BACKGROUND

[0003] Generally, for the purpose of observing cell and molecular level phenomena present in biological tissues of a live animal by in-vivo imaging, a biological microscope has been developed.

[0004] The biological microscope is characterized by in-vivo imaging, in which the imaging should be performed while the animal remains alive. Due to the characteristics of the imaging, it is very important to maintain the state of a small animal constant for the reliability of the results. Among them, the most common and important factor is the body temperature of the animal. In most in-vivo studies, an infrared heater requiring a separate installation space for maintaining the body temperature, a rigid plate heater in which body temperature feedback is not possible, or a soft silicon heater and a heating pad that are difficult to fix are used.

[0005] In addition, due to the characteristics of the biological microscope having a short penetration depth, exposing the organ to be imaged and effectively fixing the organ to not move are necessary processes in the steps before imaging. For this, compared to developing a special apparatus, manufacturing and fixing a processed product according to the respective abilities of researchers is a major approach type method.

[0006] In order to perform in-vivo imaging by a conventional handling method, a body temperature control apparatus for maintaining the body temperature of a small animal and a processed product, and a stage for fixing the small animal should be separately provided and used. Therefore, from the selection of the product to the application, a large number of repeated experiments are required according to the abilities or experiences of the respective researchers; in addition, each component is not a standardized product used separately and easily coupled to a biological microscope, so tissue exposure is performed outside the biological microscope, then the animal is moved to the stage on which a body temperature control heating pad is placed, and imaging should be performed. In this case, the fixation of the animal or the imaging tissue can be changed when the animal is moved to the stage.

[0007] In addition, when a conventional infrared heater is installed, a separate installation space is required. Meanwhile, most of the commercialized rigid plate-type heaters are of a type that maintains the temperature of the plate itself, and most of the commercialized rigid plate-type heaters do not have a function of adjusting the temperature by body temperature feedback, and most of the contents claimed as feedback have the purpose of preventing overheating to maintain the plate temperature within a predetermined range.

[0008] In addition, a silicon heater and a heating pad are designs mainly made of soft materials, and are used by being fixed to a separately prepared stage through Velcro or a tape. In the case where the heating pad is larger than the stage, the heating pad can be fixed by using Velcro or the like, in which case it is difficult to level the surface on which the small animal is placed as a whole, and the experimental animal can be placed only at a specific position, and the experimental animal is widely surrounded at the specific position of the stage, so that the position at which a glass heater or other motion correction device is fixed is limited to the stage range; whereas a heating pad or a silicon heater smaller than the stage should be fixed by a tape, and the fixed position is different each time.

[0009] Therefore, there is a need to develop a device capable of fixing the posture of a small animal and to develop a developed stage system using the device to maintain the temperature of the small animal at an appropriate level and to maintain the desired tissue in an exposed state. SUMMARY

[0010] TECHNICAL PROBLEM

[0011] The present disclosure will provide a fixing and body temperature maintenance apparatus that maintains the body temperature of a living small animal, which is an object to be observed with a biological microscope, constant.

[0012] The present disclosure will provide a fixing and body temperature maintenance apparatus that stably fixes a small animal and easily observes the small animal.

[0013] The present disclosure will provide a fixing and body temperature maintenance apparatus that, by applying a slide-type stage for smoothly moving a sample plate on which a small animal is fixed to a focusing area of a biological microscope, is capable of stably and conveniently performing high-resolution microscopic imaging while maintaining the homeostasis of various tissues of a living small animal.

[0014] According to the present disclosure, the bottom surface on which the small animal is placed is flat, and the temperature of the bottom surface itself is adjusted by body temperature feedback to maintain the body temperature of the small animal constant; and an optimized / standardized motion correction component, as can be identified in the imaging result, can be coupled to the body temperature / imaging tissue maintenance component, and can be used; and a plate heater that can be brought out and can be slid to an imaging focus position is for rapid exposure and fixation of the imaging tissue on the plate heater.

[0015] Technical solution

[0016] The fixed and body temperature maintained device according to the embodiments, for fixing a small animal to facilitate generation of a high-resolution microscopic image of a predetermined tissue of the small animal in a biological microscope having an objective lens, the device comprising: a plate heater having a heating wire mounted therein, capable of directly placing the small animal on the plate heater; a glass heater holder fixed to a hole of the plate heater; and a glass heater fixed to the glass heater holder and located above the tissue of the small animal, and maintaining the planarity of the tissue and increasing the temperature of the tissue itself.

[0017] The glass heater comprises a cover glass for adjusting the refractive index of the objective lens, a glass holder fixing the cover glass, a glass holder cover, and a heating wire heater, and

[0018] The glass holder is integrally formed with the heating wire heater, and the glass holder and the glass holder cover can be separated and fixed from each other by using screws and magnets.

[0019] The fixed and body temperature maintained device can additionally comprise: a tissue temperature sensor sensing the temperature of a biological tissue in a small animal body; a body temperature sensor sensing the body temperature of the small animal; a plate temperature sensor measuring the current temperature of the plate heater; a glass temperature sensor measuring the current temperature of the glass heater; and a control device receiving the output of the tissue temperature sensor, the output of the body temperature sensor, the output of the plate temperature sensor, and the output of the glass temperature sensor, and driving the plate heater so that the current temperature of the body temperature sensor reaches the maintenance of the target body temperature, and driving the glass heater so that the tissue temperature becomes the target tissue temperature.

[0020] Since the body temperature / homeostasis can be maintained by feedback only when there is a temperature sensor, the temperature sensor is an essential component.

[0021] A heating wire can be installed at the center of the plate heater, and equi-spaced threaded holes for installing an attachment fixing mechanism used for each tissue to be imaged can be formed on both sides of the plate heater.

[0022] A detachable rail and bar can be installed on both sides of the plate heater.

[0023] By coupling a block-type heating wire heater with the glass holder, the glass heater can be configured, and threads are provided on one side of the glass holder to fix the glass heater by using a glass heater holder, and a groove formed according to the size and thickness of the cover glass is provided on the other side of the glass holder, and a glass heater cover fixing the cover glass is covered based on a magnet or a screw to fix the cover glass to the inside of the glass holder.

[0024] The fixation and body temperature maintenance apparatus can additionally include a 3-axis stage plate, and a slide plate holder capable of detachably fixing a body temperature maintenance and fixation mechanism can be coupled to the 3-axis stage plate.

[0025] The plate heater is fixed to a slide plate holder having a rail to slide and be brought out to the outside of the stage.

[0026] Advantageous Effects

[0027] In an embodiment of the present invention, the body temperature of a live small animal, which is an object to be observed with a biological microscope, is maintained constant.

[0028] Further, in an embodiment of the present invention, the small animal is stably fixed, and the small animal is easily observed.

[0029] Further, in an embodiment of the present invention, by applying a slide stage for smoothly moving a sample plate in which the small animal is fixed to a focusing area of a biological microscope, high-resolution microscopic imaging can be stably and conveniently performed while maintaining homeostasis of various tissues of the live small animal.

[0030] Further, in the embodiment of the present application, the apparatus comprises: a device which maintains the body temperature of the small animal and exposes organs by fixing the small animal and tissues; and a stage upper plate which is capable of moving the corresponding device to a microscope focusing area to maintain homeostasis of the animal model and to more easily access internal organs and external organs, thus enhancing the user's accessibility to in vivo imaging. That is, the in vivo imaging environment of the respective existing researchers in charge is constructed in accordance with hardware / software, and thus the microscopic imaging of various tissues of the small animal can be effectively used for the research of identifying various diseases or biological phenomena existing in the animal model. BRIEF DESCRIPTION OF DRAWINGS

[0031] Figure 1 is a block diagram illustrating a fixing and body temperature maintenance apparatus according to an embodiment of the present application.

[0032] Figure 2 is a photograph illustrating a fixing and body temperature maintenance apparatus according to an embodiment of the present application.

[0033] Figure 3 is a photograph illustrating a fixing and body temperature maintenance apparatus according to an embodiment of the present application.

[0034] Figures 4 to 6 is a photograph illustrating a plate heater and a mechanism in a fixing and body temperature maintenance apparatus according to an embodiment of the present application.

[0035] Figures 7 to 9 is a photograph illustrating a plate heater and a mechanism in a fixing and body temperature maintenance apparatus according to an embodiment of the present application.

[0036] Figure 10 is a photograph in which a plate heater is coupled with a glass heater and is actually applied to imaging in a tissue living body in an abdominal cavity in a fixing and body temperature maintenance apparatus according to an embodiment of the present application.

[0037] Figure 11 is a photograph illustrating a stereotactic mount coupling for in vivo brain tissue imaging in a fixing and body temperature maintenance apparatus according to an embodiment of the present application.

[0038] Figure 12 is a photograph illustrating a stereotactic mount coupling for in vivo brain tissue imaging in a fixing and body temperature maintenance apparatus according to an embodiment of the present application.

[0039] Figure 13 is a photograph in which a rodent is fixed and imaging tissues are exposed by coupling a retractor in a fixing and body temperature maintenance apparatus according to an embodiment of the present application.

[0040] Figure 14 is a photograph showing a high-resolution / large-area imaging result of various living tissues, in a fixing and body temperature maintaining apparatus according to an embodiment of the present application, the imaging result is acquired by coupling a plate heater and a glass heater to an XYZ translation stage of a biological microscope.

[0041] Figures 15 to 18 is a photograph showing a fixing and body temperature maintaining apparatus according to an embodiment of the present application.

[0042] Figure 19 is a photograph in which, in a fixing and body temperature maintaining apparatus according to an embodiment of the present application, a slide plate holder is moved to an outermost portion in order to facilitate plate holder assembly and small animal fixing.

[0043] Figure 20 is a photograph in which, in a fixing and body temperature maintaining apparatus according to an embodiment of the present application, the plate heater on which an animal is placed is detachably fixed to the slide plate holder.

[0044] Figure 21 is a photograph in which, in a fixing and body temperature maintaining apparatus according to an embodiment of the present application, the slide plate holder on which the plate heater is fixed is moved to an imaging focus area under an objective lens. DETAILED DESCRIPTION

[0045] In the following detailed description, only certain exemplary embodiments of the application are shown and described, simply by way of illustration. As would be recognized by those skilled in the art, the present application can be modified in various different ways without departing from the spirit or scope thereof. Also, in the drawings, in order to clearly describe the present application, portions unrelated to the description have been omitted, and like reference numerals denote like components throughout the specification.

[0046] Throughout the specification, unless explicitly described to the contrary, the word "comprise" will be understood to mean the inclusion of stated components but not the exclusion of any other components. Also, the terms "-er", "-or" and "module" described in the specification mean units for processing at least one function and operation and can be implemented by hardware components or software components or combinations thereof.

[0047] Figure 1 is a block diagram showing a fixing and body temperature maintaining apparatus (of a living body of a biological microscope) according to an embodiment of the present application.

[0048] REFERENCE Figure 1, the fixing and body temperature maintaining apparatus 10 is an apparatus for fixing a small animal 300 and for fixing and body temperature maintaining alive, in order to acquire a high resolution image of an in vivo tissue of the small animal based on a biological microscope.

[0049] The fixing and body temperature maintaining apparatus 10 can be coupled to a microscope apparatus (not shown), and in order to generate a high resolution microscopic image of a predetermined tissue of the small animal 300 by using the fixing and body temperature maintaining apparatus 10, the fixing and body temperature maintaining apparatus 10 can include a glass heater 110, a plate heater 120, a glass temperature sensor 130, a tissue temperature sensor 140, a body temperature sensor 150, a plate temperature sensor 160, a glass heater holder 170, and a control device 200.

[0050] The glass heater 110 can be fixed to the glass heater holder 170, a cover glass 111 for adjusting a refractive index of an objective lens can be integrally formed with a heating wire 113, and the cover glass 111 and the heating wire 113 are located on a biological tissue of the small animal 300 to maintain flatness of the tissue and to increase a temperature of the tissue itself.

[0051] The small animal 300 can be directly placed on the plate heater 120, and a heating wire 121 is installed to control a temperature. The heating wire 121 is installed at a center of the plate heater 120 to control the temperature. Threaded holes 122 formed at equal intervals can be provided on both sides of the plate heater 120 at equal intervals, and the holes 122 are used to install an attachment type fixing mechanism used for each tissue to be imaged. A detachable rail 123 and a bar can be installed to holes 124 and can be fixed on both sides of the plate heater 120.

[0052] The fixing holes 122 of the attachment type fixing mechanism / glass heater holder and the fixing holes 124 of the rail / bar can be formed to have different specifications as needed.

[0053] The glass temperature sensor 130 senses and outputs a temperature of the glass heater 110.

[0054] The tissue temperature sensor 140 senses and outputs a temperature of a tissue.

[0055] The body temperature sensor 150 can sense and output a temperature in a rectum of the small animal 300, and use the sensed and output temperature as a body temperature.

[0056] The plate temperature sensor 160 measures and outputs a current temperature of the plate heater 120.

[0057] One side of the glass heater holder 170 can be fixed to the hole 122 of the plate heater 120, and the glass heater 110 can be fixed to the opposite side of the glass heater holder 170 by using a screw.

[0058] Referring to Figure 1 or Figure 2 The glass heater 110 can be configured by coupling a block type heating wire 113 with a glass holder 115; one side of the glass holder 115 can be fixed to the glass heater holder 170 by arranging a screw, and a groove formed according to the size and thickness of the cover glass 111 can be arranged on the other side of the glass holder; and a glass holder cover 112 fixing the cover glass 111 is covered based on a magnet 117 or a screw 116 to fix the cover glass 111 to the inside of the glass holder.

[0059] The slide type plate holder device 180 includes a slide type plate holder 181, a stage upper plate 182, and a rail 183.

[0060] The plate heater 120 and the attached fixing mechanism can be detachably coupled to the slide type plate holder 181. The slide type plate holder 181 can be coupled to the 3-axis stage upper plate 182 by using the rail 183, and the slide type plate holder 181 can be slid and brought out to the outermost portion.

[0061] The control device 200 receives an output of the glass temperature sensor 130 measuring a current temperature of the glass heater 110, an output of the tissue temperature sensor 140, an output of the body temperature sensor 150, and an output of the plate temperature sensor 160, drives the plate heater 120 so that a current temperature of the body temperature sensor 150 reaches a maintained target body temperature, and drives the glass heater 110 so that a tissue temperature becomes a target tissue temperature.

[0062] In particular, a feedback operation algorithm of software of the control device 200 is as follows.

[0063] Set parameters adjusted in the body temperature maintenance software of the control device 200 include a maintained target body temperature (set value), a maintained target temperature (set value) of the glass heater 110, and a maximum set temperature (limit) of the plate heater 120.

[0064] The control device 200 automatically controls a temperature of the plate heater 120 within a range not exceeding the maximum set temperature so that a current temperature of the body temperature sensor 150 reaches a maintained target body temperature. That is, when a temperature sensed by the body temperature sensor 150 is lower than the target body temperature, the body temperature is increased by driving the plate heater 120, and when the temperature sensed by the body temperature sensor 150 is equal to or higher than the target body temperature, the plate heater 120 is not driven.

[0065] Similarly, the control device 200 automatically controls the temperature so that the glass heater 110 in contact with the tissue sample maintains the target temperature. That is, when the temperature sensed by the tissue temperature sensor 140 is lower than the target tissue temperature, the body temperature is raised by driving the glass heater 110, and when the temperature sensed by the tissue temperature sensor 140 is equal to or higher than the target tissue temperature, the glass heater 110 is not driven.

[0066] Therefore, the glass heater 110 in contact with the upper portion of the fixed small animal 300 to be observed and the plate heater 120 in contact with the lower portion operate so as to maintain the small animal 300 at a constant temperature by the control of the control device 200 in order to acquire an image in a steady state in which the small animal 300 is maintained at a constant temperature.

[0067] The control device 200 can be one of a desktop computer, a notebook computer, or a tablet computer.

[0068] Referring to Figure 2 , the glass heater 110 is in contact with the stomach of the fixed small animal 300 to be observed, and the plate heater 120 is in contact with the back of the small animal 300. In addition, the rectal probe is manufactured so as to stably measure the body temperature of the small animal 300. During the experiment, the tissue temperature sensor 140, the body temperature sensor 150, the plate temperature sensor 160, and the glass temperature sensor 130, the plate heater 120, and the glass heater 110 can be used individually or in combination as needed.

[0069] Referring to Figure 3 , the temperature values for real-time continuous monitoring for controlling the body temperature maintenance are the current temperature of the plate heater 120, the current temperature of the body temperature sensor 150, the current temperature of the glass heater 110, the current temperature of the tissue temperature sensor (indicator 1 and indicator 2) 140, etc.

[0070] The control device 200 operates the body temperature maintenance function by executing software that monitors and controls the hardware of the plate heater 120, the hardware of the glass heater 110, the hardware of the body temperature sensor 150, and the hardware of the tissue temperature sensor (indicator 1 and indicator 2) 140, and the heaters and sensors.

[0071] Referring to Figure 3For example, the current temperature of the plate heater 120 is 38.5℃, and the limit is 40℃. The current temperature of the body temperature sensor 150 is 37℃, and the set temperature is 37℃. The current temperature of the glass heater 110 is 37.2℃, and the set temperature is 37℃. In addition, the temperature of the indicator 1 is 36.7℃, and the temperature of the indicator 2 is 36.6℃. Here, the indicators 1 and 2 as an example of the tissue temperature sensor can be implemented as one or two or more numbers.

[0072] The feedback operation algorithm of the software of the control device 200 is as follows.

[0073] The control device 200 automatically controls the temperature of the plate heater 120 within a range not exceeding the maximum set temperature, so that the current temperature of the body temperature sensor 150 reaches the maintained target body temperature. That is, when the temperature sensed by the body temperature sensor 150 is lower than the target body temperature, the body temperature is increased by driving the plate heater 120, and when the temperature sensed by the body temperature sensor 150 is equal to or higher than the target body temperature, the plate heater 120 is not driven.

[0074] The tissue temperature sensor 140 is independently used to monitor and record the temperature of a specific part of the tissue during the experiment separately from the algorithm.

[0075] Figures 4 to 6 The detailed structure of the plate heater 120 is shown in FIG. 12.

[0076] Figure 4 It is a real photo showing the plate heater, and Figure 5 It shows that the internal heating wire is disposed at the center of the plate heater 120. Figure 6 It shows the rear surface of the plate heater 120.

[0077] Referring to Figures 4 to 6 The entire plate can be heated by the heating wire 121 at the center of the plate heater 120, and equidistant threaded holes 122, detachable rails 123, and rods for mounting various attachment fixing mechanisms used for each tissue to be imaged can be installed on both sides.

[0078] Referring to Figures 7 to 10 The detailed structure of the glass heater 110 is configured by coupling the block-shaped heating wire 113 with the glass heater holder 170.

[0079] Referring to Figure 7 Threaded holes 114 are disposed at one face of the glass heater holder 170 to fix the glass heater on the face, and referring to Figure 8 or Figure 9A groove formed according to the size and thickness of the cover glass 111 is provided at the other face, and the glass heater cover 112 covering the cover glass 111 is fixed based on the magnet 117 or the screw 116 to fix the cover glass 111 inside.

[0080] Referring to Figure 10 or Figure 11 , the attachment mechanism for fixing an animal during imaging of various tissues in vivo can mainly employ a stereotaxic frame mechanism for head / brain imaging and a mechanism commonly usable for imaging of other internal organs.

[0081] First, as Figure 11 indicated, the mechanism for head / brain imaging mainly includes an ear bar 191 for fixing an ear, a tooth bar 192 as a bar for fixing a tooth, and a block 193 as a block for fixing each bar.

[0082] Referring to Figure 11 , the ear is fixed to the skull region of a small animal (300) model by the ear bar 191, and the ear bar 191 is fixed by the block 193. In addition, the protruding tooth is fixed by the tooth bar 192. Thus, in the present invention, three points are fixed to minimize the movement of the skull portion. The stereotaxic frame can be used for the purpose of fixing the head, and the body of the small animal 300 can be fixed by a separate attachment product.

[0083] Referring to Figure 12 , a result example of a high-resolution fluorescent image of an animal model acquired by actually applying the stereotaxic frame to a body temperature maintenance device for in vivo imaging of a small animal (300) is described.

[0084] Based on the fact that a CD31 antibody exhibits a fluorescent protein in immune cells in LysM-GFP mice and CX3CR1-GFP mice, the vascular endothelial cells are fluorescently stained, and a dextrain-coupled polymer fluorescent material is injected and flows inside the blood vessels to fluorescently image the blood flow.

[0085] Thus, by the fixing and body temperature maintenance device 10, the small animal 300 can be fixed and the body temperature of the small animal 300 can be maintained to acquire an in vivo high-resolution fluorescent image of the skull portion of the small animal 300 by a microscope device (not shown). In particular, both body temperature maintenance and imaging tissue fixation are performed to acquire a high-resolution fluorescent image of the skull portion.

[0086] Referring to Figure 13Mechanisms commonly used for imaging other internal organs include: a track 123 fixed to a plate heater 120, a movable rod (track module and rod) 195 mounted on the track 123 fixed to the plate heater 120, a tissue retractor 198 and a retractor fixation clamp (hook and hook clamp) 196, and a loop wire 199 and clamp 197 capable of fixing the lower body.

[0087] A retractor 198 is required to grasp the cut skin tissue to stably expose the internal organs, and the position of the retractor, including the upper and lower portions, the left and right portions, and the height, is arbitrarily controllable depending on the position of the organ to be imaged.

[0088] To achieve this, the rod 195 can be fixed to the track 123 (upper and lower portions), the retractor fixing clamp 196 can be fixed to the rod 195 (height), and the retractor 198 (left and right portions) can be fixed by using the hand screw 194.

[0089] In addition to skin tissue fixation via retractor 198, wire holder 197 can fix wire 199 via spring design so as to fix the foot by the tension of the wire, which is used for posture fixation of small animal 300.

[0090] like Figure 14 As shown, a general mouse fixation device is applied to an animal fixation and body temperature maintenance device 10 to acquire high-resolution fluorescence images of tissues in the peritoneal cavity of an animal model.

[0091] Furthermore, in the liver of a live mouse model, hepatocytes and their nuclei can be subjected to high-resolution in vivo fluorescence imaging based on fluorescent proteins, and neutrophil extracellular traps (NETs) can be subjected to high-resolution in vivo fluorescence imaging using antibody-based staining methods. In the kidneys, blood vessels can be subjected to high-resolution in vivo fluorescence imaging using fluorescent protein-based immune cells, autofluorescent kidney cells, and antibody-based staining methods. Additionally, in mouse lymph nodes, blood vessels and lymphatic vessels can be subjected to high-resolution, large-area imaging using fluorescent proteins, immune cells, and antibody-based staining methods.

[0092] Further, the embodiment of the present application generally comprises: a mechanism of a body temperature maintenance function and an attached fixation in imaging of a small animal (300) in vivo, which is directly used for exposing a tissue in order to maintain homeostasis of the small animal 300 and to achieve in vivo imaging of the small animal 300; and a slide-type plate heater 120, which can conveniently position the corresponding mechanism under an objective lens of a biological microscope.

[0093] Referring to Figures 15 to 18 , the glass heater 110 and the attached fixation mechanism are both located above the plate heater 120. By using a spring-type fixation device, the plate heater 120 can be easily attached to or detached from the slide-type plate holder device 180. A fixation groove formed according to the specification of the plate heater 120 is installed at an upper end portion of the slide-type plate holder device 180. A spring-type fixation device is installed in a diagonal direction below the left side in order to fix the spring-type fixation device to three surfaces in the groove. By using a rail 183, the slide-type plate heater holder 181 is fixed to a stage 182 and is slidable on the stage 182.

[0094] Further, a groove can be formed on the side of the rail 123 in order to easily remove the body temperature-maintained and fixed mechanism fixed to the rail 123 on the plate heater 120.

[0095] Further, Figure 19 A photograph is shown in which the slide-type plate holder 181 is moved to the outermost portion of the stage. Since the slide-type plate holder protrudes to the outside, the plate heater is easily attached / detached and the small animal is easily fixed without an obstacle such as a microscope or a lens on the upper surface of the slide-type plate holder.

[0096] In fact, Figure 20 A photograph is shown in which the plate heater 120 on which the small animal is placed is detachably fixed to the slide-type plate holder device 180.

[0097] In this state, when the plate holder is pushed and slid, a state shown in Figure 21 is shown.

[0098] Referring to Figure 21 , the slide-type plate holder 181 in which the plate heater 120 is fixed is slid and moved to an imaging focus area under the objective lens. In this state, a user can arbitrarily observe the small animal 300.

[0099] In the embodiment of the present application, the body temperature of a living small animal 300, which is an object to be observed with a biological microscope, is maintained constant.

[0100] Further, in the embodiment of the present application, the small animal 300 is stably fixed, and the small animal is easily observed.

[0101] Further, in the embodiment of the present application, by applying a slide stage for smoothly moving the sample plate in which the small animal 300 is fixed to a focusing area of a biological microscope, high-resolution microscopic imaging can be stably and conveniently performed while maintaining homeostasis of various tissues of the living small animal 300.

[0102] Further, in the embodiment of the present application, the apparatus includes a device that maintains the body temperature of the small animal 300 and exposes organs by fixing the small animal 300 and tissues, and a stage top plate that can move the corresponding device to a microscope focusing area to maintain homeostasis of an animal model and more easily access internal and external organs, thus enhancing the accessibility of a user to in vivo imaging. That is, the in vivo imaging environment that each existing researcher is in charge of is constructed in terms of hardware / software, and thus microscopic imaging of various tissues of the small animal 300 can be effectively used for research to identify various diseases or biological phenomena present in an animal model.

[0103] The embodiment of the present application is not limited to the apparatus and method described above, but can be implemented by a program for implementing a function corresponding to the configuration of the embodiment of the present application, a recording medium recording the program, etc., and those skilled in the art can easily implement the present application according to the description of the above-described embodiment.

[0104] Although the embodiment of the present application has been described in connection with a detailed description of the embodiments of the present application, it should be understood that the scope of the present application is not limited to the disclosed embodiments, but rather is intended to cover various changes, modifications and arrangements that are accessible to those skilled in the art using the basic concept of the present application as defined in the appended claims.

Claims

1. A device for immobilizing a small animal in order to generate a high-resolution microscopic image of a predetermined tissue of the small animal in a biological microscope having an objective lens, the device comprising: a plate heater having a heating wire installed therein, capable of directly placing the small animal on the plate heater; a glass heater holder fixed to a hole of the plate heater; and a glass heater fixed to the glass heater holder and located above the tissue of the small animal, and maintaining flatness of the tissue and increasing temperature of the tissue itself, wherein the glass heater comprises a cover glass for adjusting a refractive index of the objective lens, a glass holder fixing the cover glass, a glass holder cover, and a heating wire heater, and the glass holder is integrally formed with the heating wire heater, and the glass holder and the glass holder cover can be separated and fixed to each other by using a screw and a magnet.

2. The device for immobilizing and body temperature maintenance of claim 1, further comprising: a tissue temperature sensor sensing a temperature of a biological tissue in the small animal body; a body temperature sensor sensing a body temperature of the small animal; a plate temperature sensor measuring a current temperature of the plate heater; a glass temperature sensor measuring a current temperature of the glass heater; and a control device receiving an output of the tissue temperature sensor, an output of the body temperature sensor, an output of the plate temperature sensor, and an output of the glass temperature sensor, and driving the plate heater so that a current temperature of the body temperature sensor reaches maintenance of a target body temperature, and driving the glass heater so that the tissue temperature becomes a target tissue temperature.

3. The device for immobilizing and body temperature maintenance of claim 1, wherein the heating wire is installed at a center of the plate heater, and equidistantly threaded holes are formed on both sides of the plate heater for installing an attachment type immobilization mechanism used for each tissue to be imaged.

4. The device for immobilizing and body temperature maintenance of claim 1, wherein detachable rails and bars are installed on both sides of the plate heater.

5. The device for immobilizing and body temperature maintenance of claim 1, wherein the glass heater is configured by coupling a block type heating wire heater with the glass holder, and screws are provided on one side of the glass holder to fix the glass heater by using the glass heater holder, and a groove formed according to a size and thickness of the cover glass is provided on the other side of the glass holder, and a glass heater cover fixing the cover glass is covered based on a magnet or a screw to fix the cover glass to the inside of the glass holder.

6. The device for immobilizing and body temperature maintenance of claim 1, further comprising: ​ 3-axis stage top plate, a slide plate holder is coupled to the 3-axis stage top plate, and the slide plate holder can detachably fix a body temperature maintaining and fixed mechanism.

7. The fixed and body temperature maintaining apparatus of claim 6, wherein the plate heater is fixed to a slide plate holder having a rail so that the plate heater is slid and taken out to the outside of the stage.

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