Immunoassay kit for detecting M-type phospholipase A2 receptor IgG and detection method thereof

A technology of immune analysis and detection method, which is applied in the field of clinical immunology detection and can solve the problem of low activity of polypeptides

Pending Publication Date: 2020-10-16
浙江理工大学绍兴生物医药研究院有限公司 +1
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Problems solved by technology

[0004] Through further research on PLA2R, it is known that there will be migration of PLA2R antibody epitopes during the pathogenesis of IMN. Although the PLA2R antigen has ten domains, there are mainly independent epitopes in the CysR, CTLD1 and CTLD7 domains. And CysR is the main immunodominant epitope. These three epitopes will produce corresponding antibodies. Among them, the activity of the polypeptide expressed by CTLD7 alone is not high, and it must exist at the same time as CTLD6 or CTLD8. CTLD67, CTLD78, and CTLD678 will be better. activity, see figure 1

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  • Immunoassay kit for detecting M-type phospholipase A2 receptor IgG and detection method thereof
  • Immunoassay kit for detecting M-type phospholipase A2 receptor IgG and detection method thereof

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Embodiment

[0023] The preparation of described coated plate is as follows: with 50mmol / L (pH 9.6) Na 2 CO 3 -NaHCO 3 Buffer Dilute the purified recombinant protein of different epitope determinants of PLA2R (CysR or CTLD1 or CTLD67 or CTLD78 or CTLD678) to 2.5 μg / mL as coating solution, add 100 μL to each well of a 96 microwell plate, and place at 4 °C overnight; discard the coating solution, and use 50mmol / L (pH 9.6) Na 2 CO 3 -NaHCO 3 The buffer solution was washed 3 times; then each well was blocked by adding 200 μl of the above buffer solution containing 3 g / L BSA, and placed overnight at 4°C. Discard the blocking solution, pat dry and then vacuum-dry, seal the strips and store them in a freezer at -20°C until use.

[0024] The preparation of the anti-PLA2R autoantibody standard is as follows: through preliminary experiments, a serum containing high-concentration Anti-PLA2R autoantibody is screened out in patients with membranous nephropathy, and the high-concentration anti-PLA2...

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Abstract

The invention relates to an immunoassay kit for simultaneously detecting M-type phospholipase A2 receptor IgG and a detection method thereof, and belongs to the field of clinical immunological detection. The method comprises the following steps: respectively coating purified recombinant antigens of different epitope determinants of PLA2R on a microporous plate, adding diluted serum of a patient tobe detected, adding an Eu <3+>- anti-human IgG antibody, and washing and removing the unconnected Eu <3+>- anti-human IgG antibody after the labeling immune reaction is finished; and after the enhancement solution is added, respectively detecting fluorescence signals of Eu <3+> by using a time-resolved luminoscope, and obtaining the concentrations of IgG antibodies at different epitopes of the PLA2R in the sample according to the standard curve. The kit provided by the invention can be used for simultaneously detecting the contents of different epitope determinants IgG of the PLA2R, thereby providing convenience for clinical application.

Description

technical field [0001] Simultaneous detection of M-type phospholipase A2 receptor receptor (PLA2R) immunoassay kit for different epitope determinant autoantibody IgG and detection method thereof, specifically related to the detection of serum IgG content of different epitope determinant autoantibodies of PLA2R, The invention belongs to the field of clinical immunology detection. Background technique [0002] Membranous nephropathy (MN) is an autoimmune disease and a common pathological type of nephrotic syndrome, ranking first among nephrotic syndrome patients over 40 years old. About 80% of them are idiopathic membranous nephropathy (idiopathic MN, IMN) without obvious cause, and the rest are secondary MN (Secondary MN, SMN) with a cause. Common causes of SMN include: systemic lupus erythematosus, rheumatoid arthritis, hepatitis B virus infection, and drugs, poisons, tumors or environmental factors. [0003] In addition to the two necessary factors of high urinary protein...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/543G01N33/534
CPCG01N33/6854G01N33/6893G01N33/54306G01N33/534G01N2333/7056G01N2800/347
Inventor 黄飚吴青青李婷周秀梅王毅刚张丽
Owner 浙江理工大学绍兴生物医药研究院有限公司
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