Method for detecting sperm deoxyribonucleic acid fragments and film integrality
A deoxyribonucleic acid and integrity technology, applied in the field of simultaneous detection of sperm DNA fragments and membrane integrity, can solve the problem of inability to identify whether the DNA is complete, not detect the integrity of sperm DNA fragments, sperm activity, sperm membrane and DNA damage and other issues
Inactive Publication Date: 2010-12-08
SHANDONG PROVINCIAL FAMILY PLANNING INST OF SCI & TECH
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However, the simple sperm hypotonic swelling test can only determine whether a sperm membrane is complete, but cannot identify whether its DNA is complete, and it is not accurate to only detect the integrity of the sperm membrane without detecting the i...
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[0029] A method for simultaneously detecting sperm deoxyribonucleic acid fragments and membrane integrity of the present invention is characterized in that: its steps are as follows:
[0030] ① Preparation of hypotonic solution
[0031] The hypotonic solution is a solution containing 0.735g sodium citrate and 1.35g fructose per 100ml of distilled water, and the osmotic pressure is 150mOsm;
[0032] ②Preparation of eosin-Giemsa stain
[0033] Add 0.5g of eosin and 0.1g of Giemsa to every 100ml of 0.9% normal saline, mix well, make it fully dissolved, make a dye solution, and then filter with double-layer filter paper, the obtained filtrate is eosin-Gemsa dye solution, and the eosin-Giemsa staining solution was stored at room temperature at 22°C for later use;
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The invention discloses a method for detecting sperm deoxyribonucleic acid fragments and film integrality. The method comprises the following steps of: (1) preparing hypotonic solution; (2) preparing an eosin-Giemsa stain; (3) performing a simple sperm hypotonic swelling experiment; (4) performing centrifugalization on the sperm obtained by hypotonic treatment in the step (3), discarding supernate and diluting the sperm; (5) mixing the diluted sperm in the step (4) with agar gel in a certain ratio and performing incubation for 5 minutes to obtain the formulated sperm; (6) placing the formulated sperm in the step (5) on a glass slide; (7) soaking the glass slide in the solution of hydrogen chloride (HCl); (8) adding trihydroxymethylaminomethane, dithiothreitol and sulfonated succinic acid N-lauryl-based monamide disodium salt on the glass slide to soak to obtain a sample sheet; (9) washing the sample sheet in the step (8) with buffer solution; (10) dehydrating the sample sheet; and (11) drying the dehydrated sample sheet obtained by the step (10) and adding the dehydrated sample sheet into the stain for dyeing.
Description
technical field [0001] The invention relates to a detection method used in clinical medicine, specifically a method for simultaneously detecting sperm deoxyribonucleic acid fragments and membrane integrity. Background technique [0002] Sperm membrane integrity is closely related to sperm metabolism, acrosome reaction, sperm capacitation, and sperm-egg fusion. Determining whether the sperm membrane is intact can accurately reflect sperm function and predict the potential fertilization ability of sperm. Since the establishment of the sperm tail hypotonic swelling test to measure the function of human sperm membrane in 1984, the sperm tail hypotonic swelling test has become one of the few detection methods for evaluating the sperm membrane function, and it is more and more widely used in male reproduction. Basic research on contraception and clinical diagnosis and treatment of infertility. The material exchange and metabolic activities of cells rely on the cell membrane. Sper...
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Login to view more IPC IPC(8): G01N33/48G01N1/30G02B21/34C12Q1/68
Inventor 王磊光邱毅张丽红
Owner SHANDONG PROVINCIAL FAMILY PLANNING INST OF SCI & TECH
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