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Method and kit for detecting sperm acrosome activity based on electrochemistry

An acrosome enzyme and kit technology, applied in the direction of material resistance, can solve the problem of long detection time and other problems

Active Publication Date: 2019-04-05
NINGBO INST OF MATERIALS TECH & ENG CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the current detection method requires the use of complex instruments, and the detection takes a long time

Method used

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  • Method and kit for detecting sperm acrosome activity based on electrochemistry
  • Method and kit for detecting sperm acrosome activity based on electrochemistry
  • Method and kit for detecting sperm acrosome activity based on electrochemistry

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] Example 1: Detection of Human Sperm Acrosinase Using a Gold Electrode Modified with Sperm Acrosinase Substrate

[0064] 1) Gold electrode modification: Clean the gold electrode with ethanol and double-distilled water ultrasonically, and then use it for later use. figure 1 , Soak the gold electrode in 30mM cystamine dihydrochloride solution, sonicate at 50°C for 30min, dry it at room temperature, then soak the gold electrode in sperm acrosome containing 5% glutaraldehyde and 10mg / mL After incubating in the enzyme substrate PBS (0.01M, pH7.4) solution for 1h, the gold electrode was dried in the air;

[0065] 2) Gold electrode modified with sperm acrosome substrate to detect acrosome, the principle can be found in figure 2 . First establish a working curve (standard curve figure), measure the impedance G0 of the gold electrode that first measures sperm acrosome substrate modification in phosphate-buffered saline (PBSpH 7.4) (refer to image 3 A in A), prepare the solut...

Embodiment 2

[0067] Example 2: Using a gold electrode modified with a sperm acrosome substrate to detect sperm acrosome in pig sperm

[0068] 1) Gold electrode modification: Clean the gold electrode with ethanol and double-distilled water ultrasonically, and then soak the electrode in 10mL of 30mM cystamine dihydrochloride solution, sonicate at 50°C for 30min, dry it at room temperature, and then The gold electrode was soaked in a solution containing 5% glutaraldehyde and 10mg / mL sperm acrosome substrate PBS (0.01M, pH7.4) and incubated for 1h, then the gold electrode was dried in the air;

[0069] 2) Gold electrode modified with sperm acrosome substrate to detect acrosome, the principle can be found in figure 2 . First establish a working curve, measure the impedance G0 of the gold electrode modified with sperm acrosome substrate in phosphate buffered saline (PBS pH 7.4), and then prepare a series of solutions of sperm acrosome enzyme, sperm acrosome The concentration of the enzyme was...

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Abstract

The invention discloses a method for detecting sperm acrosin activity based on electrochemistry. The method comprises the steps that electrodes modified with a sperm acrosin substrate are fully soaked with sperm acrosin standard samples with different concentrations respectively, impedances of the electrodes are measured respectively, and then a standard curve of the concentration of active sperm acrosin and the electrode impedances is established; the electrodes are fully soaked with a to-be-measured sample containing sperm acrosin with an unknown concentration, impedances of the electrodes are measured, and the concentration of the sperm acrosin in the to-be-measured sample is obtained according to the standard curve, wherein the standard samples and the to-be-measured sample all adopt a buffer solution which is suitable for enabling the sperm acrosin to achieve specific binding and hydrolyzing the sperm acrosin substrate as a solvent. The invention further discloses a kit. By means of the kit and the detection method, the activity of the sperm acrosin can be detected rapidly, reliably and sensitively at a low cost, wherein detection can be completed within about 3 min, and the limit of detection can reach 0.001 micromole.

Description

technical field [0001] The invention relates to a method for evaluating the activity of sperm acrosome enzyme, in particular to a method and a kit for detecting the activity of sperm acrosome enzyme based on electrochemical technology, belonging to the field of biological detection. Background technique [0002] Acrosinase activity can reflect the quality of sperm. Insufficient acrosinase activity may lead to infertility. It is an important indicator for evaluating (modified) sperm function and fertility. Combined with routine semen analysis and sperm morphology staining (modified) Indicators can more comprehensively reflect the quality of sperm. [0003] In the detection of acrosome enzyme, radioimmunoassay or fluorescence immunoassay is often used to detect acrosome enzyme antibody in the early stage, and the fertilization ability of sperm can be judged by observing the number of acrosome enzyme in the sperm acrosome. Later, people such as Kennedy first reported the metho...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N27/04
Inventor 黄又举陈涛荣运
Owner NINGBO INST OF MATERIALS TECH & ENG CHINESE ACADEMY OF SCI
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