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Antigen for detecting sperm antibody and its expression vector, kit and detection method

A technology of expression vector and detection method, applied in the field of reproductive medicine and immune application, can solve the problems of unreliable sensitivity and accuracy, low positive rate, high false positive rate, etc., achieve reliable detection results, use less specimens, overcome sensitivity low effect

Active Publication Date: 2018-12-18
东莞兰卫医学检验实验室有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the commonly used ELISA kits in China have a high false positive rate or a low positive rate, and the sensitivity and accuracy of the domestic kits are unreliable, which cannot meet the clinical needs for infertility testing, so a new, cheap one is urgently needed. Anti-sperm antibody test kit with reliable test results to meet clinical needs

Method used

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  • Antigen for detecting sperm antibody and its expression vector, kit and detection method
  • Antigen for detecting sperm antibody and its expression vector, kit and detection method
  • Antigen for detecting sperm antibody and its expression vector, kit and detection method

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Embodiment 1

[0057] An antigen for detecting sperm antibody is a polypeptide encoded by a gene fragment of the PB functional region of the recombinant IZUMO gene, its amino acid sequence has the sequence shown in SEQ ID NO: 1, and its gene sequence has the sequence shown in SEQ ID NO: 2 .

Embodiment 2

[0059] An expression vector for detecting the antigen of sperm antibody of Example 1, the expression vector is named pET30a-IZUMO-His, and the construction diagram of this expression vector is shown in the attached figure 1 .

Embodiment 3

[0061] A method for constructing an expression vector for detecting the antigen of sperm antibody of embodiment 2, comprising the following steps:

[0062] Step 1, synthesizing the recombinant IZUMO gene PB functional region gene fragment: adopting gene synthesis to obtain the PB gene sequence and inserting a His tag at the 3' end to obtain the gene fragment having the sequence shown in SEQ ID NO:1;

[0063] Step 2, PB gene fragment amplification: using the gene fragment obtained in step 1 as a template, PCR amplification is performed to obtain the above-mentioned expression vector for detecting sperm antibody antigen.

[0064] Wherein, in step 2, the conditions of PCR amplification are: pre-denaturation at 94°C for 5 min, denaturation at 94°C for 40s, annealing at 60°C for 30s, extension at 72°C for 45s, 30 cycles, and extension at 72°C for 10 min. The reaction system is shown in Table 1.

[0065] Table 1 Recombinant IZUMO gene amplification PCR reaction system (25 μL)

[0...

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Abstract

The invention relates to the technical field of reproductive medicine and immunological application and particularly relates to an antigen for detecting a sperm antibody and its expression vector, kitand detection method. The antigen for detecting a sperm antibody is a polypeptide encoded by a PB functional region gene fragment of a recombinant IZUMO gene and has a sequence shown in the formula of SEQ ID NO: 1. The antigen for detecting a sperm antibody is a key protein of a sperm acrosome reaction and determines if the fertilization is successful. The antibody for the antigen is an importantreason leading to immune infertility. The ELISA kit for detecting the anti-sperm antibody solves the problems of low sensitivity and low specificity, has the advantages of convenience, sensitivity, few sample use amount, large-scale detection and promotion and large-scale application in the general laboratories. Therefore, the antigen is expected to be used for screening and diagnosis of anti-sperm antibodies in infertile patients.

Description

technical field [0001] The invention relates to the technical field of reproductive medicine and immunization applications, in particular to an antigen for detecting sperm antibodies, an expression vector thereof, a kit and a detection method thereof. Background technique [0002] Infertility refers to the state of couples living together for at least one year after marriage, having normal sexual life, and being unable to have children without taking any contraceptive measures. In the late 1980s, the World Health Organization (WHO) conducted a survey in 33 centers in 25 countries using standardized methods of diagnosis of infertility. Due to factors such as environment, hormones and related diseases, the number of infertile couples is increasing, and about 8% to 12% of couples worldwide have experienced infertility. Relevant studies have shown that the fertility of couples of childbearing age in my country shows a downward trend, while the infertility caused by low fertilit...

Claims

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Application Information

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IPC IPC(8): C07K14/47C12N15/70C12N15/66G01N33/68G01N33/577
CPCC07K14/47C12N15/66C12N15/70G01N33/577G01N33/68G01N2333/46
Inventor 徐军发王鑫徐建庭
Owner 东莞兰卫医学检验实验室有限公司
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