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A method for detecting sperm acrosome reaction

An acrosome reaction and sperm technology, applied in the field of sperm detection, can solve problems such as rapid detection limitations, and achieve the effect of improving efficiency

Active Publication Date: 2021-05-18
青岛复诺生物医疗有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The disadvantage of this invention is that the capacitation solution needs to be incubated for 3 hours, which has great limitations for rapid detection

Method used

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  • A method for detecting sperm acrosome reaction
  • A method for detecting sperm acrosome reaction
  • A method for detecting sperm acrosome reaction

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] A test reagent for sperm acrosome reaction, characterized in that it includes sperm capacitation treatment solution, acrosome reaction induction solution and staining solution;

[0029] The preparation method that described sperm capacitation treatment liquid contains Lactobacillus casei LC-N235 fermentation supernatant is:

[0030] (1) Take Lactobacillus casei LC-N235 and inoculate it into MRS solid medium to streak, culture at 37°C for 24 hours, pick a single colony with good growth in the plate and inoculate it into MRS liquid medium for activation, culture conditions: 32°C, 220r / min, after 24 hours of cultivation, it can be used as 1.0*10 7 (CFU / ml) Lactobacillus casei LC-N235 seed solution;

[0031] (2) Inoculate the seed liquid obtained in step (1) into the fermentation medium according to the inoculum size of 3%, and the fermentation temperature is 30°C. After 25 hours of fermentation and cultivation, centrifuge at 1000rpm for 20min, take the supernatant, filter...

Embodiment 3

[0041] The freshly taken semen was placed in a 37°C water bath for 0.5h to completely liquefy it, and then subjected to density gradient centrifugation to obtain high-motility sperm clusters to remove pollutants such as white blood cells, germ cells, and dead sperm.

[0042] Set 1 normal saline control group, 1 L-lactic acid control group (the aqueous solution of L-lactic acid of 3.5g / L), contain 3 detection groups of embodiment 1 and embodiment 2, each group is all set to 2 tubes ( for incubation at different times).

[0043] The control group was 1ml of semen diluted with normal saline, with a concentration of 1×10 6 motile sperm / ml; the detection group uses the sperm capacitation solution obtained in Example 1 and Example 2 to dilute the sperm into a concentration of 1×10 6 Motile sperm / ml suspension, each tube contains 1ml. The sperm suspensions of the control group and the test group were kept at 37°C, CO 2 Incubate in the incubator for 15 minutes and 3 hours, respecti...

Embodiment 4

[0048] Example 4 Detection of Weak Motility Sperm Motility Improvement

[0049] (1) Equilibrate the sperm capacitation solution obtained in Example 1 or Example 2 at a temperature of 37° C. for 2 hours;

[0050] (2) Put the semen collection cup with weak motility semen in a 37°C water bath for 0.5h to obtain liquefied semen;

[0051] (3) Use the preheated sperm capacitation solution obtained in Example 1 or Example 2 to wash and dilute the liquefied semen, and centrifuge at 300g for 10 minutes to obtain separated weakly motile sperm;

[0052] (4) Separate the weakly motile sperm according to 1×10 7 Add the amount of CFU / mL to the preheated sperm capacitation solution obtained in Example 1 or Example 2, and incubate at 37° C. for 120 min to obtain the incubated sperm.

[0053] (5) Set up two control groups at the same time (normal saline group with a temperature of 37°C and an L-lactic acid aqueous solution group with a concentration of 3.5g / L), wash, dilute and incubate weak...

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Abstract

The invention discloses a test reagent for sperm acrosome reaction, which is characterized in that it includes sperm capacitation treatment solution, acrosome reaction induction solution and staining solution; wherein, the sperm capacitation treatment solution contains Lactobacillus casei LC-N235 fermented supernatant. The invention also provides a kit made with the sperm acrosome reaction reagent. The invention also provides a method for detecting the acrosome by using the kit made of the sperm acrosome reaction reagent. The advantages of the present invention: (1) Breakthrough shortens the time required for sperm capacitation and improves efficiency. (2) Protect the sperm and improve the vitality of the sperm so as to reduce the deviation caused by improper preservation of the sample.

Description

technical field [0001] The invention relates to the field of sperm detection, in particular to a sperm acrosome reaction detection method suitable for high-efficiency detection by flow cytometry. Background technique [0002] The acrosome is a structure unique to sperm, and only when the sperm has a complete acrosome can the acrosome reaction (AR) occur and thus pass through the zona pellucida. Studies have shown that the percentage of acrosome reaction in live sperm has a strong positive correlation with the penetration rate of zona pellucida in sperm, and the acrosome reaction rate induced by zona pellucida has become a reliable index for evaluating sperm function. The acrosome reaction of normal fertilizing sperm must undergo the fusion and rupture of the plasma membrane and the outer acrosome membrane of the sperm head, releasing the contents, and the sign of the completion of the acrosome reaction is the complete fusion of the outer acrosome membrane. According to this ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/02C12P1/04C12R1/245
CPCC12P1/04C12N1/205C12R2001/245G01N33/5005
Inventor 吕旭楠李翼飞仪菲菲焦安军梁栋
Owner 青岛复诺生物医疗有限公司
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