JP2017167475A - Imitation ovum - Google Patents

Abstract

PROBLEM TO BE SOLVED: To increase the opportunity to learn techniques such as the cryopreservation of human ova and embryos and intracytoplasmic sperm injection.SOLUTION: The present invention provides an imitation ovum that imitates a human ovum or embryo, the imitation ovum being gelatinous and spherical with an average diameter of 100-200 μm.SELECTED DRAWING: Figure 2

Description

  The present invention relates to a simulated egg.

  Infertility is diagnosed if a person wishing to conceive cannot reach pregnancy within 1-2 years without contraception. One treatment for infertility (hereinafter referred to as infertility treatment) is in vitro fertilization (conventional in vitro fertilization, microinsemination).

  In in vitro fertilization, an embryo obtained by culturing a fertilized egg for several days (usually about 1 to 7 days) may be cryopreserved, thawed in a timely manner, and transferred to the uterus. In addition, eggs collected (before fertilization) may be frozen and stored at the request of a patient or the like. When cryopreserving human eggs (including ova and embryos), it is known that freezing with the ultra-rapid vitrification method causes less damage to eggs (almost no) compared to slow freezing. Yes. The ultra-rapid vitrification method is a method in which an egg and a small amount of cryoprotective liquid are placed on a freezing exclusive sheet, and one egg is immediately frozen in liquid nitrogen in a state of being encapsulated in a small amount of cryoprotective liquid. . Since human eggs are as small as about 150 to 200 μm, the operation of placing an egg or embryo on a freezing sheet is performed under a microscope.

  Moreover, in microinsemination (ICSI: intracytoplasmic sperm injection), sperm is injected into an ovum using a micromanipulator for cell manipulation while observing the ovum using a microscope and fertilized (see, for example, Patent Document 1).

  As described above, the above-described work accompanying in vitro fertilization is a precise work performed using a microscope or a manipulator, and thus requires skill. Conventionally, to acquire these techniques, eggs that are not suitable for embryo transfer, such as immature eggs and unfertilized eggs collected from patients, are used.

JP 2003-125750 A

  However, if eggs collected from patients are used, hospitals with a small number of patients have few opportunities for training and it is difficult to improve the technology. Therefore, there is a possibility that a difference may occur between hospitals depending on the number of patients in the level of proficiency of the above-mentioned technique that is important for success or failure in in vitro fertilization. Some people feel ethical problems in training with eggs collected from patients. Since the proficiency level of egg freezing and microinsemination techniques by ultra-rapid vitrification has a great influence on the success or failure of in vitro fertilization, it is desired to develop human resources who have mastered those techniques.

  SUMMARY An advantage of some aspects of the invention is to solve at least a part of the problems described above, and the invention can be implemented as the following forms.

(1) According to one aspect of the present invention, a simulated egg that has a spherical shape with an average diameter of 100 to 200 μm, is in a gel form, and simulates a human egg or embryo.

  Since this type of simulated egg simulates a human egg or embryo, it can be used to train work related to in vitro fertilization such as microinsemination and freezing by ultra-rapid vitrification. . Therefore, as compared with the case of using an egg collected from a patient, training opportunities can be easily increased, which contributes to the acquisition of precise techniques such as microinsemination.

(2) The simulated egg of the above form may include a spherical main body and a film that covers the entire surface of the main body. A human egg has a cytoplasm and a zona pellucida that covers the cytoplasm. In this simulated egg, the main body part simulates the cytoplasm and the membrane part simulates the zona pellucida, so that a simulated egg closer to a human egg can be provided.

(3) The simulated egg of the above form may be colored transparent or colored translucent. This facilitates identification under a microscope. Therefore, for example, it is suitable for training for beginners such as microinsemination.

(4) The main body portion of the simulated egg of the above form may be colored and transparent or colored and translucent, and the film portion may be colorless and transparent. If it does in this way, distinction between a film part and a body part will become easy.

(5) The simulated egg of the above form may further include a spherical polar body part having an average diameter of 5 to 30 μm. A human egg has a cytoplasm, a zona pellucida covering the cytoplasm, and a first polar body between the cytoplasm and the zona pellucida. In microinsemination, the first polar body may be used as an indicator of the puncture position of the injection pipette. If this simulated egg is used, training of microinsemination can be performed using the polar body as an index, and more actual microscopic insemination can be performed. Training can be performed in a state close to insemination.

(6) The simulated egg of the above form may have an average diameter of 150 to 200 μm. In in vitro fertilization, fertilized eggs that have developed into complete blastocysts and expanded blastocysts may be cryopreserved. Using this simulated egg, freezing training of complete blastocysts and expanded blastocysts can be performed.

(7) The simulated egg of the above form may contain agarose as a main component. If it does in this way, it can manufacture easily.

(8) The main body part of the simulated egg of the above form may be mainly composed of calcium alginate, and the film part may be mainly composed of agarose. In this way, a two-layer simulated egg can be easily manufactured.

(9) The simulated egg of the above-described form has an outer diameter of about 2 μm at the end of the tube, and when piercing a pipette whose tip is cut smoothly, it is elastic for 5 μm or more before the pipette enters the simulated egg. You may make it deform | transform. In this way, since the elasticity of a human egg can be simulated, training can be performed in a state closer to actual microinsemination.

(10) The simulated egg of the above form may have a density higher than that of the culture solution. In this way, since it sinks in the culture solution, the training of micro insemination can be performed easily.

(11) According to another aspect of the present invention, a simulated egg kit is provided. This simulated egg kit contains one or more simulated eggs of the above-described form and the simulated eggs together with an insoluble solution. This simulated egg kit facilitates storage and distribution.

  A plurality of constituent elements of each aspect of the present invention described above are not indispensable, and some or all of the effects described in the present specification are to be solved to solve part or all of the above-described problems. In order to achieve the above, it is possible to appropriately change, delete, replace with another new component, and partially delete the limited contents of some of the plurality of components. In order to solve part or all of the above-described problems or to achieve part or all of the effects described in this specification, technical features included in one embodiment of the present invention described above. A part or all of the technical features included in the other aspects of the present invention described above may be combined to form an independent form of the present invention.

For example, it is realizable with the manufacturing method of a simulated egg, the form of the following training apparatuses, etc.
A training apparatus for micro insemination, comprising: a microscope, a manipulator for cell manipulation, and a dish on which one or more simulated eggs taken out from the simulated egg kit of the above-described form are placed in the field of view of the microscope.
A microscope, a freezing sheet for freezing human eggs by ultra-rapid vitrification preservation method, a pipette for dropping one or more simulated eggs taken out from the simulated egg kit of the above form onto the freezing sheet, A frozen storage training device.

It is a microscope picture of the simulation egg of 1st Embodiment. It is explanatory drawing which shows typically the cross-sectional structure of a simulation egg. It is process drawing which shows the manufacturing method of a simulated egg. It is explanatory drawing which shows the mode of the training of micro insemination. It is explanatory drawing which shows the principal part of the training of micro insemination. It is explanatory drawing which shows the mode of the training of cryopreservation. It is explanatory drawing which expands and schematically shows the freezing device in which the simulated egg was mounted. It is a microscope picture of the simulated egg of 2nd Embodiment. It is explanatory drawing which shows a simulated egg typically. It is process drawing which shows the manufacturing method of a simulated egg. It is explanatory drawing which shows typically the simulated egg of 3rd Embodiment. It is a schematic diagram which shows the evaluation method of the elasticity of a simulation egg.

A. First embodiment:
FIG. 1 is a photomicrograph of the simulated egg 10 of the first embodiment. FIG. 2 is an explanatory diagram schematically showing a cross-sectional configuration of the simulated egg 10. In this specification, an egg represents the concept including an ovum (unfertilized) and an embryo (fertilized egg). The simulated egg 10 is a fine particle simulating a human egg and is a sphere having an average diameter D1 of about 150 to 160 μm. As shown in FIG. 2, the simulated egg 10 includes a main body portion 12 having a spherical shape with an average diameter D2 of about 145 to 150 μm, and a membrane portion 14 that covers the entire surface of the main body portion 12. The main body part 12 is a gel-like sphere mainly composed of calcium alginate, and the film part 14 is a thin film mainly composed of agarose having a thickness of about 5 to 10 μm. In the present embodiment, the main body portion 12 and the film portion 14 are colorless and transparent. The simulated egg 10 is in a gel form at least at 20 ° C to 40 ° C.

  In the present embodiment, the average diameter D1 of the simulated egg 10 and the average diameter D2 of the main body 12 are measured by a computer using a digital image obtained by an electron microscope and software for measuring the size of the digital image. In literatures, etc., the average diameter of a human egg is 120 ± 10 μm, and the average diameter of a blastocyst is 200 to 240 μm. When the diameter at each stage was measured (measured number of individuals n = 30), the average diameter at each stage was about 150 μm to 200 μm, so the average diameter D1 of the simulated egg 10 was about 150 to 160 μm. Here, the average diameter is an average obtained by measuring a plurality of diameters passing through the center in one egg (egg or embryo).

  FIG. 3 is a process diagram showing a method for manufacturing the simulated egg 10. First, a 1.0 w / v% sodium alginate aqueous solution 83 is sprayed by spraying into a 1.0 w / v% (weight volume percent) calcium chloride aqueous solution 82 (step S12). In step S12, a gel ball of calcium alginate is formed. However, in step S12, the diameter of the calcium alginate gel ball is not uniform. The solution in which the gel balls are formed in step S12 is filtered using a micromesh filter (step S14). Specifically, after filtration using a micromesh having a nominal size of JIS sieve (opening of a sieve) of 150 μm, the filtrate is filtered through a micromesh having a nominal size of JIS sieve of 125 μm. As a result, gel balls (main body portion 12) having an average diameter D1 of 125 μm <D1 <150 μm can be selected. The gel balls (main body portion 12) selected in step S14 are placed in a 1.0 w / v% agarose aqueous solution 85 (step S16). Heat by an autoclave at about 95 ° C. for about 30 minutes (step S18). In step S18, the agarose melts and the main body 12 does not melt. The agarose aqueous solution 88 after the process of step S18 is sprayed into the oil 87 by spraying (step S20). In step S20, the main body 12 is covered with agarose, and the simulated egg 10 is formed. In step S20, one or more main body portions 12 are covered with agarose among all the main body portions 12 included in the agarose aqueous solution 88. After the step S20, the simulated egg 10 can be obtained by filtering the oil 87 containing the simulated egg 10 using a micromesh (nominal size of JIS sieve: 150 μm) (step S22).

  FIG. 4 is an explanatory view showing a state of training for micro insemination using the simulated egg 10. FIG. 5 is an explanatory view showing the main part of the training for microinsemination. In the simulated egg 10 of FIG. 5, in order to clearly illustrate the film part 14, the film thickness is displayed thicker than the actual ratio. In the microinsemination, the worker E (for example, an embryo culture person) injects the sperm into the ovum using the microinsemination apparatus 20 using an injection pipette. In the training of micro insemination, an injection pipette is punctured into the simulated egg 10 using the micro insemination apparatus 20 that is used when actually performing micro insemination.

  As shown in FIGS. 4 and 5, the microinsemination apparatus 20 includes an inverted microscope 25, a pair of manipulators 31 and 32 for cell manipulation, and a pair of injectors 43 (the injector on the right hand side of the worker E is not shown). ), An injection pipette 34, and a holding micropipette 36. The inverted microscope 25 includes an eyepiece lens 21, a stage 22, and an objective lens. A pair of left and right manipulators 31 and 32 are disposed on the left and right sides above the stage 22. An injection pipette 34 is supported on the manipulator 31 via a holder 33 (FIG. 5). A holding micropipette 36 is supported on the manipulator 32 via a holder 35. The manipulators 31 and 32 (FIG. 4) can precisely move the injection pipette 34 and the holding micropipette 36 back and forth, right and left, and up and down, respectively, according to the operation of the operator E. An injector (not shown) is connected to the other end of the holder 33 via a flexible tube 41, and an injector 43 is connected to the other end of the holder 35 via a flexible tube 42. ing. The pair of injectors adjust the pressure in the tubes 41 and 42 and the holders 33 and 35 according to the operation of the worker E.

  When performing microinsemination training, the operator E places the petri dish 50 on the stage 22 of the inverted microscope 25 (FIG. 4). At this time, the petri dish 50 is disposed between the manipulator 31 and the manipulator 32. As shown in FIG. 5, the culture solution drop 52 formed in the petri dish 50 is covered with mineral oil 54, and the simulated egg 10 is placed in the culture solution drop 52. Since the density of the simulated egg 10 is larger than the density of the culture solution, the simulated egg 10 is submerged in the drop 52 of the culture solution. The operator E looks into the eyepiece lens 21 of the inverted microscope 25 (FIG. 4), operates the manipulators 31 and 32 while observing the inside of the petri dish 50 through the objective lens, and operates the injector 43 and an injector (not shown). Then, the simulated egg 10 is sucked and fixed to the tip of the holding micropipette 36 (FIG. 5), and the tip of the injection pipette 34 is punctured into the simulated egg 10. When the injection pipette 34 is punctured into the simulated egg 10, it is elastically deformed by about 10 μm to 15 μm before the injection pipette 34 enters the simulated egg 10. In the present embodiment, physiological saline (saline containing 0.9 w / v sodium chloride) is used as the culture solution, but is not limited thereto. When performing microinsemination training, the same culture solution as that used in actual microinsemination may be used, or an inexpensive solution may be used for training. Instead of the petri dish 50, a pallet may be used as a dish for placing the simulated egg 10 in the field of view of the inverted microscope 25. In this way, continuous training can be easily performed.

  FIG. 6 is an explanatory diagram showing the training of egg cryopreservation using the simulated egg 10. In FIG. 6, the hand of the operator E who performs the cryopreservation training by the ultra-rapid vitrification preservation method is illustrated. FIG. 7 is an explanatory diagram schematically showing an enlarged view of the freezing device 61 on which the simulated egg 10 is placed. In in vitro fertilization, an embryo obtained by culturing a fertilized egg for several days (usually about 1 to 7 days) may be cryopreserved, thawed in a timely manner, and transferred to the uterus. In addition, eggs collected (before fertilization) may be stored frozen. When cryopreserving eggs (eggs, embryos) by ultra-rapid vitrification preservation method, place an egg and a small amount of cryoprotection solution on a freezing sheet (freezing device 61), and one egg has a very small amount of cryoprotection. In a state of being encapsulated in the liquid, it is instantly frozen with liquid nitrogen. The operation of placing the egg and a small amount of cryoprotectant on the freezing device 61 is a precise operation performed under a microscope and requires skill. In this specification, the work training for placing the egg and a small amount of cryoprotectant on the freezing device 61 is referred to as “freezing preservation training”.

  In the cryopreservation training using the simulated egg 10, the operator E observes the tip of the pipette 62 and the freezing device 61 with a microscope (not shown), and applies a small amount of cryoprotective liquid from the pipette 62 to the freezing device 61. It is dripped (FIG. 6). Since the pipette 62 contains the cryoprotective liquid 70 and the plurality of simulated eggs 10, the simulated egg 10 is included in the drop of the cryoprotective liquid 70 dropped on the freezing device 61. On the freezing device 61, one simulated egg 10 is encapsulated in a small amount of cryoprotective liquid 70 (FIG. 7). In FIG. 7, in order to clearly show the cryoprotective solution 70 and the simulated egg 10, different hatchings are respectively shown. In FIG. 7, the simulated eggs 10 are included in all the cryoprotective liquid 70 drops, but the simulated eggs 10 are not included and two simulated eggs 10 are included in one drop. Sometimes.

  As described above, since the simulated egg 10 of the present embodiment is the same size as a human egg (egg, embryo), egg freezing and microinsemination by the ultra-rapid vitrification preservation method using the simulated egg 10 Can be trained. Therefore, even in hospitals where the number of patients is small, it is possible to easily increase opportunities for acquiring these techniques. Moreover, these training can also be implemented in the facility etc. outside a hospital which does not have a patient. Ethical anxiety about using eggs collected from patients for these trainings can also be resolved. As a result, it is possible to increase the number of human resources who have mastered techniques such as egg freezing and microinsemination by the ultra-rapid vitrification preservation method, and contribute to the improvement of the success rate of in vitro fertilization.

B. Second embodiment:
FIG. 8 is a photomicrograph of the simulated egg 10A of the second embodiment. FIG. 9 is an explanatory view schematically showing a simulated egg 10A. The simulated egg 10A of the second embodiment is different from the simulated egg 10 of the first embodiment in that the membrane portion 14 is not provided, that is, a single layer. The average diameter D _A 1 of the simulated egg 10A is about 150 to 160 μm.

  FIG. 10 is a process diagram showing a method of manufacturing the simulated egg 10A. First, an agarose aqueous solution 85 having a predetermined concentration is prepared (step S32). For example, an appropriate amount of 40 mL of ultrapure water and agarose are added to a 50 mL conical tube. Here, the amount of agarose necessary for preparing an agarose aqueous solution having a predetermined concentration (with respect to 40 mL of ultrapure water) is, for example, 0.12 g at 0.3 w / v% and 0.5 w / v%. 0.2 g, 0.4 g when 1.0 w / v%, 0.8 g when 2.0 w / v%, and 1.2 g when 3.0 w / v%. The agarose aqueous solution 85 prepared in step S32 is heated (for example, heated with a microwave heater) to dissolve the agarose (step S34). The agarose aqueous solution 86 in which the agarose is dissolved in step S34 is put in a pre-warmed spray, and the agarose aqueous solution 86 is sprayed into the cooled mineral oil 87 (step S36). Here, liquid paraffin may be used instead of mineral oil. In step S36, agarose gel balls are formed. In step S36, the diameter of the agarose gel ball is non-uniform. The mineral oil containing the gel balls formed in step S36 is filtered using a micromesh filter (step S38). Thereby, the simulated egg 10A can be obtained. In step S38, an agarose gel ball of a desired size may be selected with a glass pipette.

  The simulated egg 10A of the present embodiment can be used for cryopreservation training and microinsemination training. The simulated egg 10A can be manufactured more easily than the simulated egg 10 of the first embodiment.

C. Third embodiment:
FIG. 11 is an explanatory diagram schematically showing a cross-sectional configuration of the simulated egg 10B of the third embodiment. The simulated egg 10B of the third embodiment is different from the simulated egg 10 of the first embodiment in that a spherical polar body portion 16 smaller than the main body portion 12 is provided. The polar body portion 16 is disposed in the vicinity of the boundary between the main body portion 12 and the film portion 14. In the present embodiment, the average diameter _{D B} 3 polar bodies 16 is approximately 5 to 10 [mu] m. The average diameter of the polar body portion 16 is preferably about 5 to 30 μm.

  The simulated egg 10B can be manufactured by puncturing and injecting the polar body part 16 prepared in advance into the simulated egg 10 manufactured by the manufacturing method described in the first embodiment. In addition, the polar body part 16 can be manufactured by the process similar to process S12, S14 of the manufacturing method of 1st Embodiment.

  A human egg has a cytoplasm, a zona pellucida covering the cytoplasm, and a first polar body disposed between the cytoplasm and the zona pellucida. In microinsemination, the sperm is injected into the cytoplasm by inserting an injection pipette at a position far from the first polar body of the egg. The simulated egg 10B of the present embodiment includes a main body portion 12 that simulates the cytoplasm of a human egg, a membrane portion 14 that simulates a zona pellucida, and a polar body portion 16 that simulates a first polar body. In insemination training, training can be performed in a state closer to actual microinsemination.

D. Simulated egg elasticity:
The elasticity of the simulated egg was evaluated using a single-layer simulated egg similar to the simulated egg 10A of the second embodiment. As samples, the following three types of simulated eggs were prepared by changing the agarose concentration (weight percent by weight) of the agarose aqueous solution when preparing the simulated eggs.
Simulated egg 10A1: Agarose concentration 0.5w / v%
Simulated egg 10A2: agarose concentration 1.0 w / v%
Simulated egg 10A3: agarose concentration 2.0 w / v%

The evaluation method is shown below. In the following description, when the simulated eggs 10A1 to 10A3 are not distinguished, the numbers at the end of the reference numerals are omitted and referred to as simulated eggs 10A.
FIG. 12 is a schematic diagram showing a method for evaluating the elasticity of a simulated egg. 12 corresponds to a view of the petri dish 50 shown in FIG. 5 as viewed from the lower side of the drawing in FIG.

  First, a simulated egg 10A arranged in a drop 52 formed in a petri dish 50 is prepared in the same manner as in the case of microinsemination training in the first embodiment. The microinsemination apparatus 20 is operated, the simulated egg 10A is fixed by the holding micropipette 36, and the measurement injection pipette 34A is pressed against the simulated egg 10A. Here, as an injection pipette for measurement, an injection glass pipette having a diameter (outer diameter of the tube) of 2 μm and a smooth cut was used. “Smooth” means that the tip of the glass pipette is not sharp (not cut obliquely), and the central axis and the tip of the cylindrical glass pipette are substantially perpendicular. When the injection pipette 34A is pressed, the simulated egg 10A is elastically deformed and then torn (the injection pipette 34A is pierced). The amount of deformation (distance (μm)) immediately before the simulated egg 10A was broken was measured. Using the distance L (FIG. 12) between the tip of the injection pipette 34A and the outer surface of the simulated egg 10A as the amount of deformation, measured by a computer using an image captured with a microscope and software for measuring the size of the image did.

  Table 1 is a table showing the results of evaluating the elasticity of the simulated eggs. The evaluation (measurement of deformation amount) was performed using 10 each of the simulated eggs 10A1 to 10A3. The average diameter of each of the simulated eggs 10A1 to 10A3 is about 150 to 160 μm.

  As a result of the evaluation test, the simulated eggs 10A1 to 10A3 have elasticity. Since human eggs have elasticity, the use of these simulated eggs can increase the accuracy of microinsemination training. In addition, as a result of the evaluation test, it can be said that the elasticity of the simulated egg can be adjusted by adjusting the agarose concentration (w / v%) of the agarose aqueous solution when preparing the simulated egg. Therefore, by adjusting the agarose concentration of the agarose aqueous solution to produce a simulated egg having elasticity close to that of a human egg, the accuracy of microinsemination training can be further improved. The elasticity of the simulated egg is preferably 5 μm or more, more preferably 15 μm or more, based on the above evaluation method. When the amount of deformation is 15 μm or more, the elasticity of a human egg can be made closer.

E. Variations:
In addition, this invention is not restricted to said embodiment, In the range which does not deviate from the summary, it can be implemented in a various aspect, For example, the following deformation | transformation is also possible.

(1) In the above-described embodiment, the colorless and transparent simulated eggs 10, 10A, and 10B are illustrated, but may be colored and transparent or colored and translucent. By coloring, it becomes easy to visually recognize, and for example, a simulated egg suitable for training for beginners and the like can be obtained. Further, the main body 12 may be colored and transparent and colored and translucent, and the film part 14 may be colorless and transparent.

(2) In the above embodiment, the simulated eggs 10, 10A, 10B having an average diameter of about 150 to 160 μm are exemplified, but the average diameter is not limited to the above embodiment. An average diameter of 150 to 200 μm is preferable because it corresponds to the size of a human egg or embryo. An average diameter of 150 to 200 μm corresponds to the size of a human complete blastocyst and expanded blastocyst, and is suitable for training of freezing of a complete blastocyst and expanded blastocyst, for example. Moreover, you may make the average diameter of a simulated egg 100-150 micrometers. By training using a simulated egg having a size smaller than the actual size of a human egg measured by the method described in the above embodiment, the accuracy of the technique relating to in vitro fertilization can be further improved.

(3) The material which forms a simulated egg is not limited to the said embodiment, What is necessary is just a gel-like material in use condition (for example, 20 to 40 degreeC). For example, gelatin, silicon or the like can be used. Those having a density higher than those used as a culture solution, a cryoprotective solution or the like in training are preferred. During training, a solution having a density lower than that of the simulated egg may be selected as the culture solution.

(4) In the above embodiment, the example in which the polar body portion 16 is disposed in the vicinity of the boundary between the main body portion 12 and the membrane portion 14 has been described. It may be arranged closer to the center than the boundary between the film part 14 and the film part 14.

(5) The method of manufacturing the simulated egg 10 is not limited to the above embodiment, and for example, the simulated egg 10 may be manufactured using a known laminar flow principle.

(6) The simulated egg 10 may be accommodated in a container together with a solution in which the simulated egg 10 is insoluble. As a solution in which the simulated egg 10 is insoluble, physiological saline, mineral oil, a culture solution other than physiological saline, or the like may be used. The simulated egg 10 may be an insoluble solution. As the container, for example, a highly sealed container such as a disposable contact lens container can be used. This facilitates storage and distribution.

(7) In the first embodiment, the simulated egg 10 containing calcium alginate as a main component is illustrated, but agarose may be used as a main component. The main component of the main body portion 12 may be agarose, and the main component of the film portion 14 may be calcium alginate, or other gel materials such as gelatin and silicon may be used.

  The present invention is not limited to the above-described embodiments, examples, and modifications, and can be realized with various configurations without departing from the spirit thereof. For example, the technical features in the embodiments, examples, and modifications corresponding to the technical features in each embodiment described in the summary section of the invention are to solve some or all of the above-described problems, or In order to achieve part or all of the above effects, replacement or combination can be performed as appropriate. Further, if the technical feature is not described as essential in the present specification, it can be deleted as appropriate.

DESCRIPTION OF SYMBOLS 10,10A, 10B ... Simulated egg 12, 12A ... Main-body part 14 ... Membrane part 16 ... Polar body part 20 ... Microinsemination apparatus 21 ... Eyepiece lens 22 ... Stage 25 ... Inverted microscope 31, 32 ... Manipulator 33, 35 ... Holder 34 ... Injection pipette 36 ... Hold pipette 41,42 ... Tube 43 ... Injector 50 ... Petri dish 52 ... Drop 54 ... Mineral oil 61 ... Freezing device 62 ... Pipette 70 ... Cryoprotectant 82 ... Calcium chloride aqueous solution 83 ... Sodium alginate aqueous solution 87 ... oil E ... worker

Claims (11)

It has a spherical shape with an average diameter of 100 to 200 μm,
Gel-like,
Simulated eggs that mimic human eggs or embryos. The simulated egg according to claim 1,
A spherical body,
A membrane covering the entire surface of the main body,
A mock egg. The simulated egg according to claim 1 or 2,
Simulated eggs that are colored transparent or colored translucent. The simulated egg according to claim 2,
The main body is colored transparent or colored translucent,
The film part is a transparent egg that is colorless and transparent. The simulated egg according to any one of claims 1 to 3,
further,
A simulated egg having a spherical polar body part with an average diameter of 5 to 30 μm inside. The simulated egg according to any one of claims 1 to 5,
Simulated eggs having an average diameter of 150 to 200 μm. The simulated egg according to any one of claims 1 to 6,
Simulated eggs based on agarose. The simulated egg according to claim 2,
The main body is mainly composed of calcium alginate,
The membrane part is a simulated egg whose main component is agarose. The simulated egg according to any one of claims 1 to 8,
A simulated egg having an outer diameter of about 2 μm at the end of the tube and elastically deforming by 5 μm or more before the pipette enters the simulated egg when puncturing a pipette whose end is cut smoothly. The simulated egg according to any one of claims 1 to 9,
Simulated eggs with a density higher than that of the culture solution.   A simulated egg kit in which one or more simulated eggs according to any one of claims 1 to 10 are contained in a container together with the insoluble solution.