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Flow detection reagent for human anti-Mullerian hormone, preparation method of flow detection reagent and application

An anti-Müllerian hormone and flow detection technology is applied in the field of flow detection reagents for human anti-Müllerian hormone and its preparation, which can solve the problem of large fluctuations in laboratory detection values ​​and no unified AMH detection standard calibration measurement. questions of value

Active Publication Date: 2017-07-21
ZHEJIANG CELLPRO BIOTECH
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  • Claims
  • Application Information

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Problems solved by technology

However, with the extensive and in-depth application of AMH, higher requirements are put forward for its laboratory testing, and there are still some urgent problems to be solved: (1) There is no unified AMH testing standard and calibration value; (2) Due to the detection technology, the detection values ​​of each laboratory fluctuate greatly; this provides the research and development direction and application prospect for the new ANH detection technology

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  • Flow detection reagent for human anti-Mullerian hormone, preparation method of flow detection reagent and application
  • Flow detection reagent for human anti-Mullerian hormone, preparation method of flow detection reagent and application

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specific Embodiment 2

[0047] The preparation method of the flow detection reagent of human anti-Müllerian hormone, the specific steps are as follows:

[0048] (1) Preparation of AMH monoclonal antibody

[0049] First, obtain the complete coding sequence of AMH protein and the amino acid sequence of the translated mature corresponding protein from the Uniprot website, and use protein-related software to analyze the stability of the amino acid sequence, epitope, and the structure and sequence of AMH under physiological conditions; select A mature AMH protein sequence (the mature fragment is relatively stable and will not be degraded) (the amino acid sequence of the mature AMH protein fragment is as follows: SAGATAADGPCALRELSVDLRAERSVLIPETYQANNCQGVCGWPQSDRNPRYGNHVVLLLKMQARGAALARPPCCCVPTAYAGKLLISLSEERISAHHVPNMVATECGCR) as the antigen for preparing the anti-Müllerian hormone antibody library, and the base sequence of the antigen was cloned On the prokaryotic expression vector pET-41a, the E.coli prokary...

specific Embodiment 3

[0065] For the application of the above-mentioned flow detection reagent for human anti-Mullerian hormone, the specific steps of using the flow detection reagent to detect the content of human anti-Müllerian hormone are as follows: add the human serum or blood of the sample to be tested to the antibody Ab1- Add a certain volume of antibody Ab2-fluorescent molecular marker solution to the microsphere conjugate solution, and incubate at 37 degrees for 30 minutes. INHB in the sample combines with Ab1 antibody microsphere conjugate and Ab2 antibody fluorescent molecular marker to form a microsphere Ball-Ab1-AMH-Ab2-fluorescent molecule complex, the fluorescence emitted by the fluorescent molecule can be detected by flow cytometry, and the content of human anti-Müllerian hormone in the sample is calculated according to the relationship between the fluorescence intensity and the concentration of AMH , wherein the molar ratio of antibody Ab1-microsphere conjugate to antibody Ab2-fluor...

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Abstract

The invention discloses a flow detection reagent for human anti-Mullerian hormone, a preparation method of the flow detection reagent and application. The reagent is characterized by comprising an antibody Ab1-microsphere conjugate used for detecting the content of anti-Mullerian hormone in human serum or blood, and an antibody Ab2-fluorescent molecular marker. The preparation method comprises the following steps: preparing and pairing human anti-Mullerian hormone monoclonal antibodies, coupling between the human anti-Mullerian hormone monoclonal antibodies and carrier microspheres, and fluorescent molecular marking of the monoclonal antibodies. The flow detection reagent has the advantages that the content of AMH is directly judged according to fluorescence values, an experimental result can be obtained only by adding a blood sample into the reagent, the experimental operation steps during detection are simplified, moreover, high-throughput and multi-sample continuous detection can be realized through an automatic sample adding system, the detection efficiency is greatly improved, and the detection sensitivity can reach 0.01ng / ml.

Description

technical field [0001] The invention relates to a detection technology for the content of anti-Müllerian hormone in serum or plasma, in particular to a flow detection reagent for human anti-Müllerian hormone and its preparation method and application. Background technique [0002] Anti-Mullerian hormone (AMH) is a member of the transforming growth factor β superfamily, first discovered by Professor Alfred Jost. AMH is made of two identical 70×10 3 Da subunit, a dimeric glycoprotein connected by disulfide bonds, with a relative molecular mass of 140×10 3 Da; The human AMH coding gene is located on the short arm of chromosome 19, with a size of 2.4-2.8 kb and 5 exons. Anti-Müllerian hormone (AMH) plays an important role in the development of gonad organs and is one of the important markers of gonad function in men and women. In males, AMH is mainly produced by Leydig cells of the testis, starting from embryogenesis and throughout life; during the development of male fetuses...

Claims

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Application Information

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IPC IPC(8): G01N21/64G01N33/577
CPCG01N21/6486G01N33/577
Inventor 高攀房海燕陈继勇
Owner ZHEJIANG CELLPRO BIOTECH
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