Vinyl pack for experimental animal

Abstract

To provide a method for biological imaging such as MRI imaging while maintaining a microorganism state of an experimental animal.SOLUTION: A vinyl pack for an experimental animal that has a predetermined configuration and a method for biological imaging of an experimental animal using the same are provided.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] The present invention relates to a vinyl pack for laboratory animals. [Background technology]

[0002] Magnetic resonance imaging (MRI) and other bioimaging techniques are widely used as a means of non-invasively evaluating the structure and function of the body, and are applied to laboratory animals in medical and biological research. Meanwhile, microbiologically controlled germ-free animals, gnotobiotic animals, specific pathogen-free (SPF) animals, and infected animals are widely used as laboratory animals. Germ-free animals are animals raised in a sterile environment and are free of microorganisms. Gnotobiotic animals are animals in which all microorganisms present are known. Gnotobiotic animals are generally created by colonizing specific microorganisms in germ-free animals. SPF animals are animals that are free of specific microorganisms, parasites, and other pathogens, but are not necessarily free of other microorganisms. Infectious animals are animals that have been infected with pathogens.

[0003] Germ-free animals are generally kept in a sterile environment using vinyl isolators. To perform MRI imaging on germ-free animals, the animals must be removed from the sterile environment and placed in an MRI machine. However, because MRI machines are large and expensive, it is difficult for research facilities to possess MRI machines dedicated to germ-free animals. Therefore, these machines are typically used for measuring non-sterile specimens as well. This makes it difficult to maintain the entire MRI machine in a sterile state. Furthermore, it is practically impossible to make the entire MRI measurement room sterile. Therefore, when performing MRI imaging on germ-free animals, the only option is to place the germ-free animals, which have been kept in a sterile environment, in a non-sterile MRI machine. After MRI imaging, the sterile state of the experimental animals is lost, making the animals unsuitable for subsequent germ-free animal experiments.

[0004] Therefore, in previous experiments, the only way to perform MRI imaging on germ-free animals was to increase the size of the starting animal population. For example, in this method, a large population of animals (n) is reared in a sterile environment in a vinyl isolator. From this, a first subpopulation (n1) is removed for the first round of MRI imaging, the remaining (n-n1) individuals are continued to be reared in a sterile environment in a vinyl isolator, a second subpopulation (n2) is removed for the second round of MRI imaging, and the remaining (n-n1-n2) individuals are continued to be reared in a sterile environment in a vinyl isolator. Similarly, to perform multiple MRI imaging rounds (rounds 3, 4, and m), the population n must be continuously increased. However, this method results in increased research costs and animal resources as the population (n) grows.

[0005] In addition, in the case of rats and mice, it was possible to maintain a certain degree of genetic uniformity in the experimental animal population by constructing the parent population n from inbred strains or cloned individuals. However, in the case of marmosets and macaques, there are no inbred strains, and it is difficult to produce cloned individuals. Therefore, the offspring population n1 measured in the first round and the offspring population n measured in the final round were m Therefore, in MRI imaging of germ-free animals, the MRI results of offspring population n1 measured in the first round and offspring population n measured in the final round were different for genetically non-uniform individuals. m The same applies to gnotobiotic and spf animals.

[0006] Non-Patent Document 1 reports on the basic experimental techniques for non-invasive small animal MRI and MRS. The experimental animals are small animals such as mice, which are not germ-free. Furthermore, the environment in which MRI imaging is performed is not a sterile environment.

[0007] Although there have been reports of MRI measurements of germ-free mice, in all cases the mice were euthanized during the MRI measurement or only MRIs of individual mice were reported after death (Non-Patent Documents 2 and 3).To the inventors' knowledge, there have been no reports of germ-free experimental animals undergoing MRI measurements while still alive, then being returned to a germ-free environment for rearing, and then undergoing another MRI measurement after a certain period of time.

[0008] Patent document 1 describes a vinyl isolator for animal surgery. Patent document 2 describes an oxygen-controlled environment device for surgical procedures, transplants, and wound healing. Patent document 3 describes a sterilized enclosure. Patent document 4 describes an aseptic work system. These documents are not related to bioimaging such as MRI imaging.

[0009] There is a need for a method for performing MRI imaging on the same experimental animal while maintaining its microbial state. There is also a need for a method for performing MRI measurements on germ-free animals while maintaining their germ-free state without euthanizing them, and then returning the germ-free animals to a sterile breeding environment. There is also a need for a method for performing MRI imaging of infected animals without contaminating the environment of the imaging equipment while ensuring the safety of the workers. [Prior art documents] [Patent documents]

[0010] [Patent Document 1] Patent Publication No. 2021-108772 [Patent Document 2] US Patent Application Publication No. 2002 / 0045796 [Patent Document 3] International Publication No. 2005 / 092229 Brochure [Patent Document 4] Japanese Patent Application Publication No. 2019-115324 [Non-patent literature]

[0011] [Non-Patent Document 1] J. Vis. Exp. (32), e1592, DOI:10.3791 / 1592 (2009) [Non-patent document 2] PLos One 13(8),e0201829,DOI:10.1371 / journal.pone.0201829 (2018) [Non-patent document 3] Cell Host Microbe 29(7), 1199-1208.e5,DOI: 10.1016 / j.chom.2021.05.002 (2021) Summary of the Invention [Problem to be solved by the invention]

[0012] The present invention aims to at least partially solve the above problems. For example, the present invention aims to provide a method for performing MRI imaging of laboratory animals while maintaining their microbial state. Another example of the present invention aims to provide an inexpensive and easy-to-use means for bioimaging of laboratory animals. [Means for solving the problem]

[0013] The inventors have thoroughly investigated the above problems and demonstrated that a vinyl pack having a specific configuration enables bioimaging of laboratory animals, including germ-free animals, and have completed the present invention, which encompasses these embodiments.

[0014] The present invention includes the following embodiments. [1] A plastic pack for laboratory animals having at least a first tracheal tube, a second tracheal tube, and a pipe for connecting the laboratory animal to a vital sign monitoring device. [2] A vinyl pack for laboratory animals according to embodiment 1, which is made of a transparent, soft material that can be sealed and closed. [3] A plastic pack for laboratory animals according to embodiment 1 or 2, having at least one closed end and at least one open end. [4] A vinyl pack for laboratory animals according to embodiment 3, wherein the vinyl pack for laboratory animals has a truncated cone shape and the outer diameter of the closed end is smaller than the outer diameter of the open end. [5] A vinyl pack for laboratory animals according to embodiment 1, wherein the vital sign monitoring device is a pulse oximeter. [6] A plastic pack for laboratory animals according to embodiment 1, wherein the pipe can be connected to a rubber stopper through which a cable of a vital sign monitoring device is passed. [7] A system for bioimaging of laboratory animals, in which the vinyl pack for laboratory animals described in embodiment 1 or 6 is connected to a vital sign monitoring device. [8] A system for bioimaging of laboratory animals, in which the tracheal tube described in embodiment 1 is connected to a ventilator via a filter. [9] In a vinyl isolator (i) anesthetizing the experimental animal; (ii) intubating one end of the first tracheal tube into the experimental animal; (iii) connecting the other end of the first tracheal tube to a ventilator via a filter; (iv) placing the experimental animal in a vinyl pack for experimental animals according to any one of embodiments 1 to 6; (v) connecting a vital signs monitoring device to the laboratory animal via a pipe; and (vi) sealing the vinyl pack for laboratory animals according to any one of embodiments 1 to 6 with a heat sealer or sealing clip placed inside the vinyl isolator while maintaining the microbial condition of the laboratory animal; A method for preparing an experimental animal for performing in vivo imaging, comprising:

[10] (vii) placing the experimental animal in the plastic pack prepared by the method of embodiment 9 in an imaging device of one of the following groups; a) MRI, b) CT scan, c) X-rays, d) Ultrasound imaging; or e) in vivo fluorescence imaging; and (viii) imaging the positioned experimental animal; An imaging method comprising:

[11] A method for returning a laboratory animal in a vinyl pack after imaging to a re-transportation vinyl isolator while maintaining the microbial state, the re-transportation vinyl isolator having an incision film, (ix) A process of adhering the vinyl pack containing the experimental animal to the incision film of the re-transportation vinyl isolator using double-sided adhesive tape; (x) a step of simultaneously cutting the incision film of the re-transportation vinyl isolator and the vinyl pack for the laboratory animals; (xi) A process of transferring the laboratory animals contained in the vinyl packs for laboratory animals into the vinyl isolator for re-transportation; The method comprising:

[12] (xii) transferring the experimental animals from the re-transportation vinyl isolator described in embodiment 11 to a breeding vinyl isolator; (xiii) a step of rearing laboratory animals in a vinyl isolator; performing the method according to embodiments 9 and 10 on the laboratory animals; and Optionally repeating steps (xii) and (xiii) and the method according to embodiments 9 and 10, A method for imaging the same individual experimental animal multiple times while maintaining the microbial state, comprising: [Effects of the Invention]

[0015] The present disclosure allows for biological imaging to be performed while maintaining the microbial state of laboratory animals. [Brief explanation of the drawings]

[0016] [Figure 1]This is a photograph of a plastic pack 100 for laboratory animals. The plastic pack shown is frusto-conical in shape. The closed end 5 on the left has a diameter (outer diameter) of approximately 35 mm. The open end 6 is on the right. The overall longitudinal length of the plastic pack is approximately 700 mm. These shapes and dimensions are for illustrative purposes only. [Figure 2] 1 is a photograph of the vinyl pack 100 for experimental animals when the experimental animal 4 (marmoset) is packed therein. [Figure 3] This shows an overview of the technology that allows MRI imaging of germ-free marmosets while maintaining their sterile condition in vivo. In the first stage shown in the upper left, the experimental animal 4 is anesthetized in the transport vinyl isolator 20, packed, and then transported from the transport vinyl isolator 20. In the second stage shown in the center, the experimental animal 4 is transported, placed in the MRI device 70, and MRI imaging is performed. In the third stage shown in the lower right, the experimental animal 4 is re-transported into the transport vinyl isolator 40 and allowed to wake up. [Figure 4] This is a photograph of the equipment used. From left to right, there is an anesthesia machine 30, a transport cart 50, a ventilator 55, a vinyl isolator for transport 20, and a vinyl isolator for rearing 60. The configuration is an example. [Figure 5] This is a photograph of the tracheal intubation of experimental animal 4 (germ-free marmoset) in a vinyl isolator 20. [Figure 6] This is a photograph of the packaging of a laboratory animal 4 (a germ-free marmoset) in a vinyl pack 100 for laboratory animals. The open end 6 of the vinyl pack 100 for laboratory animals is sealed using a heat sealer 28. This operation is performed in a sterile environment. [Figure 7] 1 is a photograph of a transport cart 50 for transporting a packed experimental animal 4 (a germ-free marmoset). [Figure 8] This is a photograph showing a packed experimental animal 4 (germ-free marmoset) being placed in an MRI apparatus 70. [Figure 9]This is a photograph of the operation of reloading experimental animal 4 (germ-free marmoset) into reload vinyl isolator 40. This photograph is of the operation of adhering a vinyl pack with double-sided adhesive tape 44 to incision film 42 placed in reload vinyl isolator 40. [Figure 10] This is a photograph of the procedure for reintroducing experimental animal 4 (germ-free marmoset) into the vinyl isolator. This photograph shows the film and vinyl pack being incised at the same time. [Figure 11] This is a photograph of the procedure for re-transporting experimental animal 4 (germ-free marmoset) into the vinyl isolator. This photograph shows the animal being placed in the vinyl isolator and awakened. [Figure 12] 1 is a schematic diagram showing the re-transportation operation of the laboratory animal 4 (germ-free marmoset) into the re-transportation vinyl isolator 40. The double-sided adhesive tape 44 between the incision film 42 and the laboratory animal vinyl pack 100 is not shown. [Figure 13] This is an MRI image of the marmoset's head. [Figure 14] This is a photograph of a CT scan of an experimental animal using a vinyl pack 100 for experimental animals. [Figure 15] This is a photograph showing in vivo fluorescence imaging of an experimental animal using a vinyl pack 100 for experimental animals. DETAILED DESCRIPTION OF THE INVENTION

[0017] The present invention will now be described in detail with reference to the drawings.

[0018] In one embodiment, the present invention provides a vinyl pack 100 for laboratory animals. Examples of laboratory animals include, but are not limited to, germ-free animals, gnotobiotic animals, SPF animals, and infected animals. Infectious animals refer to animals infected with a pathogen, such as germ-free animals, gnotobiotic animals, or conventional laboratory animals infected with a known or unknown virus. Conventional laboratory animals refer to laboratory animals that are not germ-free, gnotobiotic, or SPF animals, but are not raised in a special environment such as a sterile environment. When the vinyl pack 100 for laboratory animals of the present disclosure is used for germ-free animals, it may be referred to as a vinyl pack for germ-free animals. When the vinyl pack 100 for laboratory animals of the present disclosure is used for gnotobiotic animals, it may be referred to as a vinyl pack for gnotobiotic animals. When the vinyl pack 100 for laboratory animals of the present disclosure is used for infected animals, it may be referred to as a vinyl pack for infected animals. When the vinyl pack 100 for laboratory animals of the present disclosure is used for SPF animals, it may be referred to as a vinyl pack for SPF animals. In this specification, the vinyl pack for laboratory animals may be referred to as the vinyl pack for animal bioimaging, but these are synonymous.

[0019] The laboratory animal vinyl pack 100 of the present invention has at least two tracheal tubes. In one embodiment, the laboratory animal vinyl pack 100 of the present invention has two tracheal tubes. For convenience, these will be referred to as a first tracheal tube 1, a second tracheal tube 2, ..., an nth tracheal tube. The laboratory animal vinyl pack 100 of the present invention has at least one pipe 3. In one embodiment, the laboratory animal vinyl pack 100 of the present invention has one pipe 3. The pipe 3 is for connecting a vital sign monitoring device to the animal.

[0020] The laboratory animal vinyl pack 100 of the present invention is made of a transparent, flexible material. Examples of the transparent, flexible material include vinyl, such as vinyl chloride. In one embodiment, the transparent, flexible material used in the vinyl pack can be the same vinyl used in vinyl isolators, such as vinyl chloride. Vinyl chloride membranes are flexible, have high elongation, and are generally transparent or highly transparent. Furthermore, the transparent, flexible material used in the vinyl pack can be bonded by high-frequency electromagnetic waves and can be easily handled. In one embodiment, the transparent, flexible material used in the vinyl pack can be, but is not limited to, a vinyl film having a thickness of 0.05 to 1 mm, 0.08 to 1 mm, 0.3 to 1 cm, for example, 0.4 to 0.8 cm, for example, 0.4 to 0.6 cm, or for example, 0.5 cm to 0.55 cm. Known or commercially available vinyl films can be used, including, but not limited to, those manufactured by Achilles. The transparent, flexible material can be sealed or closed with a sealing clip.

[0021] An example of a laboratory animal vinyl pack 100 of the present invention is shown in FIG. 1. The left side has a closed end 5, and the right side has an open end 6. For ease of operation, the diameter (outer diameter) of the open end 6 can be larger than the diameter of the closed end 5. This allows operations such as inserting a tracheal tube into the laboratory animal to be performed near the right open end 6, where there is more space, and the laboratory animal can then be moved toward the closed end 5 within the vinyl pack. Therefore, in one embodiment, the laboratory animal vinyl pack 100 can be shaped like a truncated cone. In another embodiment, the laboratory animal vinyl pack 100 can be shaped like a cylinder. In this case, the diameter of the open end 6 and the diameter of the closed end 5 can be the same. The laboratory animal vinyl pack 100 can also be shaped like a truncated triangular pyramid, a truncated square pyramid, or another similar shape.

[0022] In one embodiment, the first tracheal tube 1 can be used for supplying anesthetic gas. In another embodiment, the first tracheal tube 1 can be used for supplying and exhausting air for artificial respiration. In another embodiment, the second tracheal tube 2 can be used for exhausting (inhaling). Unless otherwise specified, the tracheal tube is connected to a filter 7 to keep the inside of the experimental animal vinyl pack 100 sterile. The tracheal tube can also be connected to an artificial respirator 55.

[0023] In another embodiment, the experimental animal vinyl pack 100 may have first, second, and third tracheal tubes. In this case, the first tracheal tube 1 may be used for supplying anesthetic gas, the second tracheal tube 2 may be used for exhaust, and the third tracheal tube may be used for supplying air for artificial respiration.

[0024] Pipe 3 may have any shape as long as it can connect vital sign monitoring device 11 to laboratory animal 4 inside the pack. In one embodiment, pipe 3 for connecting to vital sign monitoring device 11 is a cylindrical pipe. In one embodiment, pipe 3 is made of polyvinyl chloride.

[0025] For convenience, the midpoint from the closed end 5 to the open end 6 in the longitudinal direction of the vinyl pack 100 for laboratory animals is referred to as the center of the vinyl pack 100 for laboratory animals. In one embodiment, the first tracheal tube 1, the second tracheal tube 2, and the pipe 3 may be positioned at the center, near the center, or closer to the open end 6 than the center of the vinyl pack 100 for laboratory animals. In one embodiment, the pipe 3 may be positioned at or near the center of the vinyl pack 100 for laboratory animals, and the first tracheal tube 1 and the second tracheal tube 2 may be positioned closer to the open end 6 than the center of the vinyl pack 100 for laboratory animals. In one embodiment, the first tracheal tube 1 and the second tracheal tube 2 may be positioned at or near the center of the vinyl pack 100 for laboratory animals, and the pipe 3 may be positioned closer to the open end 6 than the center of the vinyl pack 100 for laboratory animals.

[0026] In certain embodiments, the longitudinal length of the vinyl pack for laboratory animals 100 may be 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, for example, 100% or more longer than the length of the laboratory animal 4 to be housed. In certain embodiments, the longitudinal length of the vinyl pack for laboratory animals 100 may be a length that allows the laboratory animal 4 to be housed at the center of the vinyl pack for laboratory animals 100 or between the center and the closed end 5. In certain embodiments, the longitudinal length of the vinyl pack for laboratory animals 100 may be about 1.5 times or more, about 1.8 times or more, for example, about 2 times or more the length of the laboratory animal 4 to be housed. The length of the laboratory animal 4 here refers to the length from the top of the head to the tips of the toes when the laboratory animal 4 is housed in the closed end of the vinyl pack for laboratory animals 100.

[0027] Examples of tracheal tubes include, but are not limited to, known tracheal tubes and commercially available tracheal tubes, such as Atom Pink Catheter (Atom Medical). The tracheal tube can be crimped into a vinyl pack 100 for laboratory animals. Crimping can be performed using the following procedure: First, a hole is made in the vinyl pack. Alternatively, a vinyl pack with holes is prepared. The tracheal tube is passed through the hole. In one embodiment, the vinyl pack has one hole, and two tracheal tubes are passed through it. In another embodiment, the vinyl pack has two holes, and one tracheal tube is passed through each hole. The same procedure can be used when using three or more tracheal tubes. A piece of vinyl chloride is placed on the vinyl pack over the portion where the tracheal tube protrudes, and the piece is crimped while applying heat (seam bonding). Crimping can be performed using, for example, a high-frequency welder or a high-frequency sewing machine. The high-frequency welder can be a known or commercially available device. Examples include Seidensha Electronics' KV-3000 series, such as the KV-3000T, the KV-4000 series, and the KV-5000 series, but are not limited to these.

[0028] The vital sign monitoring device 11 may be any device that monitors vital signs. Examples of vital signs include, but are not limited to, respiration, body temperature, electrocardiogram, and arterial oxygen saturation. The vital sign monitoring device 11 may monitor one or more of these. Examples of vital sign monitoring devices 11 include, but are not limited to, pulse oximeters, thermometers, electrocardiogram monitoring devices, and respiration monitoring devices. Respiration can also be monitored by measuring the concentration of CO2 or anesthetic gases. In one embodiment, the vital sign monitoring device 11 is a pulse oximeter.

[0029] The cable 9 for the vital signs monitoring device may be passed directly through the pipe 3, or the cable 9 for the vital signs monitoring device may be passed through a sealing means such as a rubber stopper 8, which is then connected to the pipe 3.

[0030] FIG. 3 shows an overview of a technology that can perform MRI imaging on a laboratory animal 4 (a germ-free marmoset) while maintaining its in vivo germ-free state. In the first step, the laboratory animal is anesthetized and packed in a vinyl isolator, and the germ-free animal is removed from the vinyl isolator. For convenience, the vinyl isolator used for the packing operation is sometimes referred to as the removal vinyl isolator 20 in this specification. First, the laboratory animal 4 is anesthetized in the removal vinyl isolator 20. Anesthesia can be administered by injection or by supplying anesthetic gas. Next, the laboratory animal 4 is intubated with a tracheal tube 1 for artificial respiration. See, for example, FIG. 5 . A vital sign monitoring device 11 is also connected to the laboratory animal 4. For example, a pulse oximeter can be attached to the laboratory animal. For example, the pulse oximeter cable can be passed through a rubber stopper 8, which can be connected to a pipe 3. The laboratory animal 4 is housed in the closed end 5 of the laboratory animal vinyl pack 100. In one embodiment, the head of the laboratory animal 4 is housed in the closed end 5 of the laboratory animal vinyl pack 100. This has the advantage that the head of the experimental animal 4 can be easily placed in the MRI device 70 when performing MRI imaging of the brain. In another embodiment, the feet of the experimental animal 4 are placed on the closed end 5 side of the experimental animal vinyl pack 100. This has the advantage that the feet of the experimental animal 4 can be easily placed in the MRI device 70 when measuring the shape of the experimental animal's limbs, etc. Next, the open end 6 of the experimental animal vinyl pack 100 is sealed. This can be done using a heat sealer 28, a laminator, or a sealing clip. This series of operations is performed in a sterile environment within the export vinyl isolator 20. Both tracheal tubes can be connected to filters 7. This allows the interior of the experimental animal vinyl pack 100 to maintain a sterile environment. Next, the experimental animal vinyl pack 100 containing the experimental animal 4 is removed from the export vinyl isolator 20. This removal can be performed, for example, via a steril lock 25. Therefore, the export vinyl isolator 20 can have a steril lock 25.

[0031] In the second stage, the packed experimental animal 4 is transported and placed in the MRI apparatus 70, where MRI imaging is performed. For example, if the output vinyl isolator 20 where the packing process is performed and the MRI apparatus 70 are in different rooms, a transport means such as a transport cart 50 may be used for transportation. FIG. 7 shows an example of the transport cart 50. In one embodiment, the transport cart 50 may have a platform for carrying the vital sign monitoring device 11, the ventilator 55, and the experimental animal vinyl pack 100. Next, the experimental animal vinyl pack 100 containing the experimental animal 4 is placed in the MRI apparatus 70. An example is shown in FIG. 8.

[0032] After the MRI scan is completed, the third step involves returning the experimental animal 4 to the vinyl isolator and allowing it to awaken. To maintain a sterile environment for the experimental animal 4, a vinyl isolator with an incision film 42 may be used. For convenience, this vinyl isolator may be referred to herein as a re-entry vinyl isolator 40. In one embodiment, the incision film 42 may be placed at the bottom of the re-entry vinyl isolator 40, but this is not limitative. The experimental animal vinyl pack 100 may be attached to the incision film 42 with double-sided adhesive tape 44. See, for example, FIG. 9 . A set of sterilized instruments, including a scalpel, may be placed inside the re-entry vinyl isolator 40. Next, from inside the re-entry vinyl isolator 40, the incision film 42 and the experimental animal vinyl pack 100 may be incised simultaneously with a scalpel using operating gloves or the like. See, for example, FIG. 10 . Next, the experimental animal 4 is re-entered from the experimental animal vinyl pack 100 into the re-entry vinyl isolator 40. See, for example, FIG. 12 . In a specific embodiment, the cut double-sided adhesive tape 44 and the laboratory animal vinyl pack 100 can be clipped together with tweezers from inside the re-entry vinyl isolator 40, thereby maintaining a sterile environment within the re-entry vinyl isolator 40. In another embodiment, the clipping operation may be omitted. The laboratory animal 4 is then re-entered from the opened laboratory animal vinyl pack 100 into the re-entry vinyl isolator 40. Once the laboratory animal 4 has been transferred into the re-entry vinyl isolator 40, the laboratory animal 4 may be awakened. Alternatively, the laboratory animal 4 may be further transferred from the re-entry vinyl isolator 40 to the breeding vinyl isolator 60.

[0033] In one embodiment, the output vinyl isolator 20 and the re-entry vinyl isolator 40 may be a common vinyl isolator. The output vinyl isolator or the re-entry vinyl isolator may be referred to herein as a treatment vinyl isolator.

[0034] The double-sided adhesive tape 44 used in the re-carrying procedure may be, for example, transparent soft vinyl chloride (e.g., 0.02 mm thick, manufactured by Nippon Wave Block Co., Ltd.) having adhesive or bonding materials on both sides, but is not limited to this. The double-sided adhesive film, for example, transparent soft vinyl chloride, may have a thickness of 0.01 mm to 1 mm, 0.02 mm to 0.5 mm, for example, 0.03 mm, for example, 0.02 mm, or for example, 0.01 mm.

[0035] In certain embodiments, the reloading vinyl isolator 40 may have one or more observation windows. The observation windows may be made of a hard window material, such as transparent hard vinyl chloride (a thin plate with a thickness of 0.1 mm). This reduces diffuse reflection of light and shortens the time required for incision and reloading. In certain embodiments, a platform for placing the experimental animal may be used to place the experimental animal in close contact with the bottom of the reloading vinyl isolator 40. See, for example, FIG. 9.

[0036] The unloading vinyl isolator 20 and the reloading vinyl isolator 40 can be fitted with one or more, two or more, for example, three, four, five, six, seven, or eight gloves, but are not limited to this. The unloading vinyl isolator 20 and the reloading vinyl isolator 40 can have the usual configuration of a vinyl isolator, for example, one or more opening and closing parts for connecting to the steril lock 25, an intake part for supplying sterile air, and an exhaust part for discharging air.

[0037] In one embodiment, the present invention provides a bioimaging system. The bioimaging system includes at least a plastic bag for laboratory animals. In one embodiment, the bioimaging system further includes a vital sign monitoring device. In one embodiment, the vital sign monitoring device is a pulse oximeter, and therefore the bioimaging system further includes a pulse oximeter. In one embodiment, the vital sign monitoring device can be connected to the laboratory animal. In one embodiment, the pulse oximeter can be connected to the laboratory animal. In one embodiment, the pulse oximeter cable can be connected to the laboratory animal by passing it through a rubber stopper. The rubber stopper can be passed through a pipe 3. In one embodiment, the bioimaging system includes a ventilator. In this case, the first tracheal tube 1 can be connected to the ventilator via a filter. The bioimaging system can further include an MRI device. The bioimaging system can further include an export vinyl isolator. A heat sealer or a sealing clip can be placed inside the export vinyl isolator. The bioimaging system can further include a re-import vinyl isolator. The re-import vinyl isolator can have an incision film. The re-import vinyl isolator can be connected to the breeding vinyl isolator.The biological imaging system of the present invention allows MRI imaging to be performed multiple times on the same individual laboratory animal.

[0038] In some embodiments, the experimental animal 4 may be a vertebrate. Examples of vertebrates include mammals and birds. Mammals include marmosets, mice (including nude mice), rats, hamsters, guinea pigs, ferrets, rabbits, dogs, cats, pigs (including minipigs and microminipigs), rhesus monkeys, cynomolgus monkeys, chimpanzees, gorillas, etc. Birds include, but are not limited to, chickens and quails. In certain embodiments, the experimental animal 4 may be a young animal, for example, 1 week, 2 weeks, 3 weeks, 4 weeks, 1 month, 2 months, 3 months, 4 months, 5 months, or 6 months old. Unless otherwise specified, the term "animal" used herein excludes humans. In other words, non-human animals are simply referred to as "animals" in this specification. In some embodiments, the experimental animal 4 may be a microbiologically controlled animal or an animal infected with a specific pathogenic microorganism. In some embodiments, the experimental animal 4 may be a gnotobiotic animal or an SPF animal.

[0039] The present invention makes it possible to evaluate changes in MRI imaging over time for laboratory animals, particularly the same individual laboratory animal. For example, a germ-free animal can be reared in a sterile environment, placed in a laboratory animal vinyl pack of the present invention, subjected to MRI imaging, and then returned to a vinyl isolator for rearing. After rearing for a certain period of time, the same individual can be subjected to multiple MRI measurements. To the inventors' knowledge, this has not been possible with conventional methods, particularly for germ-free marmosets.

[0040] The present invention allows multiple MRI measurements to be performed on the same experimental animal without euthanizing it, thereby reducing the number of experimental animals that need to be euthanized, which is advantageous in line with the 3R philosophy of animal testing.

[0041] Furthermore, the present invention can contribute to research on intestinal bacteria. For example, a relationship between the intestinal microbiota and diseases such as Parkinson's disease, Alzheimer's disease, and obesity has been pointed out. The present invention enables comparisons that were previously impossible. Specifically, by first performing MRI imaging on a germ-free animal, then transplanting a certain microbiota into the germ-free animal, and then performing MRI imaging on the animal after the microbiota transplant, it becomes possible to compare the images before and after the transplant.

[0042] For convenience, this specification has mainly described an MRI device and MRI imaging, but the effects of the present invention can be similarly obtained when CT scans, in vivo fluorescent imaging, X-ray photography, or ultrasound imaging are performed instead of MRI imaging. Therefore, this specification encompasses embodiments in which "MRI device" is replaced with "CT scan device," "in vivo fluorescent imaging device," "X-ray photography," or "ultrasound imaging," and in which "MRI imaging" is replaced with "CT scan," "in vivo fluorescent imaging," "X-ray photography," or "ultrasound imaging."

[0043] The present invention makes it possible to reduce the number of animals in accordance with the 3Rs of laboratory animals and obtain experimental results that are not affected by individual differences.

[0044] (Example) An example of MRI imaging of a germ-free marmoset is described below. Four germ-free marmosets, two males and two females, were used, maintained in a vinyl isolator. Figure 2 shows a germ-free marmoset packed in a vinyl bag for laboratory animals. A tracheal catheter (Atom Pink Catheter 6Fr, Atom Medical) crimped onto the vinyl bag for laboratory animals was inserted into the marmoset's trachea. Two 0.22 μm filters (Millex Syringe Filter, Merck) were attached to the outer end of the catheter to prevent microbial contamination. A pulse oximeter probe (Nellcor Oxysensor III, Covidien Japan) was attached to the marmoset's lower leg, passing through the rubber stopper in the pipe of the vinyl bag for laboratory animals. Similar procedures can be used with various laboratory animals, such as mice and rats.

[0045] For MRI imaging, the breeding vinyl isolator and the carry-in / out vinyl isolator were connected, and the marmoset, which had been given anesthesia by injection in the breeding vinyl isolator, was moved to the carry-in / out vinyl isolator as shown in Figure 4. After that, the marmoset, which had been given anesthesia by injection in the carry-out isolator, was intubated, and a pulse oximeter probe was attached to its lower leg, and the rear end of the experimental animal vinyl pack was sealed by crimping it with a heat sealer as shown in Figure 6.

[0046] The packed marmoset was removed from the transport / import vinyl isolator via a sterilized lock, placed on a transport cart, and connected to a ventilator (SN-480-7, Shinano Seisakusho) for anesthesia maintenance with isoflurane. It was then transported to the MRI room while undergoing biomonitoring with a pulse oximeter. After transport, the packed marmoset was transferred to the MRI bed, as shown in Figure 8, and its head was fixed in a helmet-type receiving coil. MRI measurements were performed using a Bruker Biospec 7T-MRI, and brain images were acquired. Figure 13 shows an example of an MRI image. Anesthesia was maintained in the same manner during the MRI, and biomonitoring was also performed. A CT scan was performed using a Rigaku CosmoScan FX using the same procedure, as shown in Figure 14. In vivo fluorescence imaging was also performed using a PerkinElmer IVIS Lumina, as shown in Figure 15.

[0047] After MRI imaging, the packed marmoset was transferred to a transport cart and transported to the breeding room. One side of the vinyl pack for experimental animals containing the marmoset was then attached with double-sided adhesive tape to the incision film of the vinyl isolator for transporting / unloading, as shown in Figure 9. As shown in Figures 10 and 11, the attached film and the vinyl pack were simultaneously incised from inside the vinyl isolator for transporting / unloading, and the animal was introduced into the vinyl isolator through the incision hole. After waking up, the animal was returned to the vinyl isolator for breeding.

[0048] Sterility tests were performed on each animal using a culture method 3-4 weeks after MRI imaging. Feces, feed, and VI swabs were used as specimens and cultured in thioglycollate medium at room temperature and 37°C, and in potato dextrose medium at room temperature for 2 weeks each. All sterility tests resulted in negative results, with no viable bacteria or fungi detected, demonstrating that MRI imaging can be performed on germ-free marmosets in vivo while maintaining a germ-free state using the method using vinyl laboratory animal packs.

[0049] Furthermore, in this embodiment, the cases of MRI imaging, CT scan, and in vivo fluorescence imaging device have been described, but the present invention is not limited to these, and PET (positron emission tomography), X-ray imaging, and ultrasound imaging diagnosis can also be performed using similar procedures.

[0050] In this example, a heat sealer was used as an example of sealing the vinyl pack for laboratory animals, but sealing can also be performed using a sealing clip.

[0051] Furthermore, although this example describes an example of imaging germ-free animals, the present invention can maintain animals in a state where the movement of microorganisms is blocked inside and outside the vinyl pack, so that biological imaging can be performed not only on germ-free animals, but also on gnotobiotic animals, SPF animals, and infected animals, maintaining the microbial state and preventing the spread of microorganisms.

[0052] Furthermore, although the present embodiment has been described with reference to MRI imaging, the present invention is not limited to MRI imaging, and CT scans, X-ray imaging, and ultrasound image diagnosis can be performed using similar procedures.

[0053] As another embodiment of the present invention, the same procedure as in the first embodiment can be carried out using various experimental animals such as mice and rats. [Industrial Applicability]

[0054] According to the present disclosure, imaging such as MRI can be performed multiple times on the same experimental animal while maintaining the microbial state of the animal.

[0055] Numerous documents, including manufacturer's manuals, are cited herein. The disclosures of these documents, while not considered relevant to the patentability of this invention, are hereby incorporated by reference in their entirety. More particularly, all referenced documents are hereby incorporated by reference to the same extent as if each individual document was specifically and individually indicated to be incorporated by reference. [Explanation of symbols]

[0056] 1. First tracheal tube 2 Second tracheal tube 3 Pipe 4. Experimental animals 5 Closed end 6 open end 7 Filters 8 Rubber stopper (sealing means) 9 Cable 10 Crimping section 11. Vital Signs Monitoring Devices 20 Vinyl isolator for transport 25 Sterilrock 28 Heat Sealer 30 Anesthesia Machine 40 Re-entry vinyl isolator 42 incision film 44 double-sided adhesive tape 50 Transport Cart 55 Respirator 60 Vinyl Isolator for Raising Animals 70 MRI machine 100 Vinyl Bags for Laboratory Animals

Claims

1. A plastic pack for laboratory animals having at least a first tracheal tube, a second tracheal tube, and a pipe for connecting the laboratory animal to a vital sign monitoring device.

2. 2. The vinyl pack for laboratory animals according to claim 1, which is made of a transparent, soft material that can be sealed and closed.

3. 3. The plastic pack for laboratory animals according to claim 1 or 2, which has at least one closed end and at least one open end.

4. 4. The plastic pack for laboratory animals according to claim 3, wherein the plastic pack for laboratory animals has a truncated cone shape and the outer diameter of the closed end is smaller than the outer diameter of the open end.

5. 2. The plastic pack for laboratory animals according to claim 1, wherein the vital sign monitoring device is a pulse oximeter.

6. 2. The plastic pack for laboratory animals according to claim 1, wherein the pipe can be connected to a rubber stopper through which a cable of a vital sign monitoring device is passed.

7. A system for bioimaging of laboratory animals, comprising the vinyl pack for laboratory animals according to claim 1 or 6 connected to a vital sign monitoring device.

8. A system for bioimaging of laboratory animals, comprising the tracheal tube according to claim 1 connected to an artificial respirator via a filter.

9. Inside a vinyl isolator (i) anesthetizing the experimental animal; (ii) intubating one end of the first tracheal tube into the experimental animal; (iii) connecting the other end of the first tracheal tube to a ventilator via a filter; (iv) placing a laboratory animal in the vinyl pack for laboratory animals according to any one of claims 1 to 6; (v) connecting a vital signs monitoring device to the laboratory animal via a pipe; and (vi) sealing the vinyl pack for laboratory animals according to any one of claims 1 to 6 with a heat sealer or sealing clip placed inside the vinyl isolator while maintaining the microbial condition of the laboratory animal; A method for preparing an experimental animal for performing in vivo imaging, comprising:

10. (vii) placing the experimental animal in the plastic pack prepared by the method of claim 9 in an imaging device of one of the following groups; a) MRI, b) CT scan, c) X-rays, d) Ultrasound imaging; or e) in vivo fluorescence imaging; and (viii) imaging the positioned experimental animal; An imaging method comprising:

11. A method for returning a laboratory animal in a vinyl pack after imaging to a re-transportation vinyl isolator while maintaining the microbial state, the method comprising: (ix) a step of adhering the experimental animal vinyl pack containing the experimental animal to the incision film of the re-transportation vinyl isolator with double-sided adhesive tape; (x) a step of simultaneously cutting the incision film of the re-transportation vinyl isolator and the vinyl pack for laboratory animals; (xi) transferring the laboratory animals contained in the vinyl packs for laboratory animals into a vinyl isolator for re-import; The method comprising:

12. (xii) transferring the laboratory animal from the re-transportation vinyl isolator according to claim 11 to a breeding vinyl isolator; (xiii) rearing laboratory animals in a vinyl isolator; carrying out the method according to claims 9 and 10 on the bred laboratory animals; and Optionally repeating steps (xii) and (xiii) and the methods of claims 9 and 10, A method for imaging the same individual experimental animal multiple times while maintaining the microbial state, comprising:

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