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Sperm DNA fragment detection kit and detecting method thereof

A detection kit and kit technology, applied in the field of biochemical detection, can solve the problems of poor color rendering effect of sperm tail and need to improve the accuracy rate, and achieve the effect of improving the quality of sperm staining, wide application range, and clear boundaries

Inactive Publication Date: 2015-10-21
CHENGDU PUHUA TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The SCD preservation solution mentioned in this application can effectively preserve the semen specimen, so that the DNA fragmentation rate does not change within two weeks of freezing at -20°C, which is beneficial to the preservation of the specimen; No, the counting accuracy needs to be improved

Method used

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  • Sperm DNA fragment detection kit and detecting method thereof
  • Sperm DNA fragment detection kit and detecting method thereof
  • Sperm DNA fragment detection kit and detecting method thereof

Examples

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preparation example Construction

[0053] 1. Preparation of sperm DNA fragment detection kit

[0054] Liquid Ⅰ: containing 0.04~0.08mol / L hydrochloric acid;

[0055] II solution: containing 7.305~146.1g / L sodium chloride, 1.86~37.2g / L ethylenediaminetetraacetic acid, 0.0607~1.214g / L tris, 0.5~20g / L lauryl sarcosine sodium nitrite, 1-20mL / L polyethylene glycol octylphenyl ether, 4.32-86.4mL / L 3-mercapto-1,2-propanediol, solution pH=7-7.5;

[0056] 0.5-2% agarose-coated glass slides;

[0057] 0.5-2% low melting point agarose;

[0058] Cover slip; Wright's stain; Wright's buffer;

[0059] Wherein "%" represents the weight percent concentration.

[0060] The preparation method of each composition of the above kit is as follows:

[0061] Preparation method of liquid Ⅰ:

[0062] 1) Take 30-100mL of pure water and put it in a 1000mL volumetric flask, measure 0.335-6.7mL of concentrated hydrochloric acid, add it into the volumetric flask, and stir to mix.

[0063] 2) Dilute to 1000mL with pure water, stir and mix....

Embodiment 1

[0093] This example provides a sperm DNA fragment detection kit, including:

[0094] Liquid Ⅰ: containing 0.08mol / L hydrochloric acid;

[0095] Solution II: containing 2.5mol / L sodium chloride, 0.1mol / L ethylenediaminetetraacetic acid, 10mmol / L trishydroxymethylaminomethane, 2% sodium lauryl sarcosine, 2% polyethylene glycol octane phenyl ether, 0.8mol / L 3-mercapto-1,2-propanediol, solution pH=7;

[0096] 1% agarose-coated slides;

[0097] 1% low melting point agarose;

[0098] Cover slip; Wright's stain; Wright's buffer;

[0099] Wherein "%" represents the weight percent concentration.

[0100] Preparation method of liquid Ⅰ:

[0101] 1) Take 30-100mL of pure water and put it in a 1000mL volumetric flask, measure 6.7mL of concentrated hydrochloric acid, add it into the volumetric flask, stir and mix well.

[0102] 2) Dilute to 1000mL with pure water, stir and mix.

[0103] Preparation method of liquid II:

[0104] 1) Take 30-100mL of pure water and place it in a 1000m...

Embodiment 2

[0133] select with figure 1 Same sample, add appropriate amount of hydrogen peroxide in the detection process, adopt the same method detection with embodiment one, obtain as follows figure 2 Microscopic results shown. As shown in the figure, it can be seen that after adding hydrogen peroxide, the halo of the sperm disappeared with this kit, indicating that the DNA of the sperm was damaged.

[0134] The specific method of adding hydrogen peroxide is: in the detection method step 2) the fresh semen sample is diluted to 10~20×10 6 After cells / mL, add 100 μL of hydrogen peroxide solution, incubate at 37° C. for 10 minutes, and then perform step 3) agarose melting and subsequent incubation.

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Abstract

The invention discloses a sperm DNA fragment detection kit and a detecting method thereof. The kit comprises mixed liquid of liquid I, hydrochloric acid, liquid II, sodium chloride, ethylenediamine tetraacetic acid, trihydroxymethyl aminomethane, sarcosyl, polyethylene glycol octylphenol ether and 3-sulfydryl-1, 2-propylene glycol, and further comprises a glass slide covered with agarose, low-gelling temperature agarose, Wright's stain, Wright's buffer solutions and the like. The detecting result reliability, sperm staining quality and the like of the kit are greatly improved. An image of the detecting result of the kit is clear, formed halos are clear, a boundary of the halos and a head is obvious, and the sperm DNA injury degree can be judged conveniently. According to the kit and the detecting method of the kit, standardization of a laboratory is facilitated, analysis costs are low, no special instrument or reagents are needed, the application range is wide, the detecting needs of small and medium laboratories are met, and the kit and the method of the kit can be suitable for detecting an DNA fragment of a single sperm. The kit can be compatible with a later automatic instrument, and is used for detecting the large laboratories, and detecting efficiency is improved.

Description

technical field [0001] The invention belongs to the technical field of biochemical detection, in particular to the technical field of DNA detection, in particular to a sperm DNA fragment detection kit and a detection method thereof. Background technique [0002] The integrity of genetic material is crucial for successful pregnancy and the health of offspring. The degree of DNA fragmentation can reflect the integrity of sperm genetic material, in-depth evaluation of male fertility, prediction of treatment outcome and guidance of treatment. At present, the commonly used methods for detecting sperm DNA fragments mainly include: sperm chromatin structure analysis test, single-cell gel electrophoresis test, terminal transferase-mediated dUTP end-labeling method, fluorescence in situ hybridization technology, and 8-hydroxydeoxyguanosine determination. Wait. However, most of these methods require flow cytometry or fluorescence microscope analysis and observation, or the detection ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCG01N33/5005
Inventor 许文明
Owner CHENGDU PUHUA TECH CO LTD
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