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Cryopreservation liquid and cryopreservation method for oocytes

A cryopreservation liquid and oocyte technology, applied in the preservation, application, and animal husbandry of human or animal bodies, can solve the problems of lack of oocyte freezing system, improve survival rate and developmental potential, and reduce freezing damage and cryotoxicity, subsequent fertilization and embryonic development

Active Publication Date: 2020-02-28
JINAN CENTER HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In view of the lack of a freezing system for oocyte characteristics in the prior art, the present invention provides an oocyte cryopreservation solution and preservation method to solve the above technical problems

Method used

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  • Cryopreservation liquid and cryopreservation method for oocytes

Examples

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Effect test

Embodiment 1

[0026] A cryopreservation solution for oocytes, including a balance solution 1, a balance solution 2 and a vitrification solution, the balance solution 1 includes base solution, 2.5% ethylene glycol, 1.25% dimethyl sulfoxide , 1.75% propylene glycol.

[0027] The second equilibrium solution includes base solution, 7.5% ethylene glycol, 3.75% dimethyl sulfoxide, 3.75% propylene glycol, and 10 μM (μmol / L) genistein.

[0028] The vitrification solution includes base solution, 15% ethylene glycol, 7.5% dimethyl sulfoxide, 7.5% propylene glycol, 0.5M (mol / L) sucrose, and 10 μM (μmol / L) genistein.

[0029] The base solution is a base reagent containing 20% ​​SPS (serum protein substitute) by mass fraction.

[0030] The basic reagent is Earle's balanced salt solution.

[0031] A method for cryopreservation of oocytes, which adopts the concentration progressive method to freeze, comprising the following steps: the oocytes are first placed in the balance solution 1 for 1-2 minutes, t...

Embodiment 2

[0033] A cryopreservation solution for oocytes, including a balance solution 1, a balance solution 2 and a vitrification solution, and 1 part of the base solution;

[0034] The balance liquid one includes base liquid, 2.5% ethylene glycol, 1.25% dimethyl sulfoxide, and 1.75% propylene glycol.

[0035] The second equilibrium solution includes base solution, 7.5% ethylene glycol, 3.75% dimethyl sulfoxide, 3.75% propylene glycol, and 10 μM (μmol / L) genistein.

[0036] The vitrification solution includes base solution, 15% ethylene glycol, 7.5% dimethyl sulfoxide, 7.5% propylene glycol, 0.5M (mol / L) sucrose, and 10 μM (μmol / L) genistein.

[0037] The base solution is a base reagent containing 20% ​​SPS (serum protein substitute) by mass fraction.

[0038] The basic reagent is Earle's balanced salt solution.

[0039] A method for cryopreservation of oocytes, which adopts the concentration progressive method to freeze, comprising the following steps: the oocytes are first placed i...

experiment example 1

[0041] Mouse embryos and discarded human oocytes were used for freezing experiments, and the specific experimental methods were as follows.

[0042] 1. Freezing:

[0043] Take this oocyte cryopreservation solution out of the refrigerator and return it to room temperature, equilibrate at room temperature for more than 20 minutes, make 0.5ml droplets in a petri dish, and transfer 2-4 oocytes that need to be frozen to the base solution Put it in the medium for 1 minute, transfer it to the balance solution 1 and let it stand for 1-2 minutes, then transfer it to the balance solution 2, carefully observe the shrinkage and recruitment of the oocyte, usually after shrinkage, it will recover to about 80% of the original volume, and wait for The cell membrane can be restored to smoothness (about 6-8 minutes), then move the oocyte to the vitrification solution for 30 seconds to 1 minute, transfer it to the marked Cryotop carrier rod, and quickly put it into liquid nitrogen. Set the oute...

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Abstract

The invention belongs to the technical field of oocyte freezing, and particularly relates to a cryopreservation liquid and a cryopreservation method for oocytes. The cryopreservation liquid comprisesone part of an equilibrium liquid I, one part of an equilibrium liquid II and one part of one vitrification liquid. According to the invention, focused on the cell structure of oocytes, a specific freezing reagent for oocytes is innovatively designed by optimizing a conventional freezing system, improving the proportions of freezing components and adding a specific freezing protective agent; and through verification via mouse embryo and waste human oocyte freezing experiments, the specific freezing reagent for oocytes is proved to have greatly improved effects compared with existing embryo freezing reagents.

Description

technical field [0001] The invention belongs to the technical field of oocyte freezing, and in particular relates to an oocyte cryopreservation solution and a preservation method. Background technique [0002] Oocyte cryopreservation can preserve the fertility of women with ovarian function decline caused by age and medical factors, which is conducive to improving the oocyte donation process and alleviating a series of problems caused by embryo cryopreservation. At present, oocyte freezing mainly adopts vitrification technology, which can significantly reduce the cell damage caused by ice crystal formation when oocytes are frozen. However, the developmental potential of oocytes after cryopreservation is still significantly lower than that of fresh oocytes. oocyte. Studies have shown that drastic changes in temperature and osmotic pressure during freezing and the toxicity of cryoprotectants will cause damage to the cytoskeleton, microtubules and spindles, increasing the rate...

Claims

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Application Information

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IPC IPC(8): A01N1/02
CPCA01N1/0221
Inventor 张迎春武斌田姗胡瑞路西兰卞凤娇
Owner JINAN CENTER HOSPITAL
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