External detection method for sperm-oocyte interaction

A technology for in vitro detection of oocytes, applied in biological testing, material inspection products, fluorescence/phosphorescence, etc., to improve detection efficiency, reduce detection costs, and reduce operating time

Inactive Publication Date: 2009-07-15
刘瑜
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] The technical problem to be solved by the present invention is to reduce the consumption...

Method used

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Embodiment Construction

[0015] The present invention is described in further detail below.

[0016] a sample

[0017] Sperm can be obtained from fresh liquefied semen, or thawed sperm that can be cryopreserved in liquid nitrogen.

[0018] Oocytes are fresh oocytes or oocytes stored at 2-8°C in egg cell preservation solution. The oocytes stored in egg cell preservation solution were washed 3 times with culture medium before use.

[0019] Second reagent

[0020] 1. Culture medium

[0021] The culture medium is used to extract high-motility sperm, wash live gametes, and provide energy and suitable reaction conditions for sperm and oocytes.

[0022] The weight content of NaCl in the culture solution of the present invention is 0.28-1.11%, the weight content of KCl is 0.02-0.07%, and the weight content of CaCl 2 2H 2 O weight content is 0.01-0.05%, KH 2 PO 4 The weight content is 0.01-0.03%, MgSO4·7H 2 O weight content is 0.01-0.06%, phenol red weight content is 0.0005-0.003‰, NaHCO 3 The weight...

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Abstract

The invention provides a sperm-oocyte interaction vitro detection method, comprising: a. extracting sperms by a swim-up method or density gradient method; b. mixing the sperms and oocyte extracted from the step a according a certain proportion in the culture medium and setting a blank control of the oocyte in the culture medium; c. removing the sperms which are bonded with the surface of the oocyte in the step b and detecting the number of sperms bonded with the transparent area of the oocyte under an inverted phase contrast microscope; d. separating the oocyte from the sperms bonded with the transparent area of the oocyte and respectively collecting the separated sperms and the oocyte; e. coating the sperms in the blank control of the oocyte and sperms separated by the step b on the glass slide, drying and fixing the sperms using the fixation fluid, cleaning the fixed sperms and adding fluorescein-agglutinin label, re-cleaning the sperms after reaction, detecting the sperm acrosome reaction rate under a fluorescence microscope; f. detecting the number of sperms penetrating into the oocyte separated by the step d under the inverted phase contrast microscope. The detection method reduces the consumption of the label resource and the detection cost and time and increases the detection efficiency.

Description

technical field [0001] The present invention relates to a method for detecting sperm-oocyte interaction in vitro, more specifically, a method for jointly detecting sperm-oocyte interaction in vitro. Background technique [0002] Mammalian sperm must have good physiological functions to complete the fertilization process. Fertilization is a complicated process, which needs to go through the process of combining sperm with the zona pellucida, acrosome reaction induced by the zona pellucida, and sperm passing through the zona pellucida and combining with the vitelline membrane. Any problem in any link can lead to fertilization failure. These processes can be evaluated in vitro by sperm-oocyte interaction assays to find the etiology of infertility and provide a direct scientific basis for the selection of assisted reproductive methods. [0003] Sperm-oocyte interaction tests include sperm-zona pellucida binding test, zona pellucida-induced sperm acrosome reaction test, and sper...

Claims

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Application Information

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IPC IPC(8): G01N33/48G01N21/64
Inventor 刘瑜
Owner 刘瑜
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