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Human body anti-AQP4 autoantibodies detection material and preparation method thereof

A technology for detecting materials and autoantibodies, applied in the field of biomedicine, can solve the problems of low sensitivity and specificity of the CBA method, difficult to clearly interpret the detection results, and complicated operation process, so as to increase the detection sensitivity and specificity, and interpret the detection results. Easy to identify, reduce the effect of non-specific signal

Active Publication Date: 2017-08-08
SHAANXI MYBIOTECH CO LTD
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Problems solved by technology

However, the main drawbacks of this type of detection method are: (1) Various proteins in the cell may cause background coloring problems
In the specific detection process, although there are idling control cells for comparison, when the expression of exogenous antigens is low, some detection results are still difficult to clearly interpret
(2) The operation process is complicated, and the accuracy of each step directly affects the test results, which is very dependent on the experience of the monitors. In addition, when performing immunofluorescence or flow cytometry experiments, special instruments are required, such as fluorescence microscopes or flow cytometers
The sensitivity and specificity of the IIF method is relatively lower than that of the CBA method, which may be caused by the differences between species in the reaction substrates.

Method used

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  • Human body anti-AQP4 autoantibodies detection material and preparation method thereof
  • Human body anti-AQP4 autoantibodies detection material and preparation method thereof

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Embodiment Construction

[0031] The present invention will be further described in detail below in conjunction with specific embodiments.

[0032] The preparation method of the human anti-AQP4 autoantibody detection material provided by the present invention specifically comprises the following steps:

[0033] 1. Artificially synthesized or obtained AQP4 full-length target gene by PCR method, construct HEK293 / pCI-neo-M1-AQP4, HEK293 / pCI-neo-M23-AQP4 cells stably or transiently expressing M1 and M23 subtypes, and empty them HEK293 / pCI-neo cells were used as the control group. The specific steps are as follows:

[0034] 1. Plasmid construction

[0035] 1.1 Obtaining the target gene: obtain the full-length target gene of AQP4 of M1 and M23 subtypes by artificial synthesis or PCR method, and add NotI / NheI enzyme cutting sites at both ends.

[0036] 1.2 Insert the obtained two target genes into the vector pCI-neo plasmid respectively, the insertion site is NotI / NheI, name the obtained two plasmids respe...

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Abstract

The invention provides a human body anti-AQP4 autoantibodies detection material and a preparation method thereof. The preparation method comprises the steps of firstly acquiring M1 and M23 subtype AQP4 overall length target genes, dyeing into an HEK293 cell, and building an HEK293 / pCI-neo-M1-AQP4 cell and an HEK293 / pCI-neo-M23-AQP4 cell expressing M1 and M23 subtype AQP4; extracting an AQP4-cell membrane complex of the cells; finally curing the AQP4-cell membrane complex on a carrier, and obtaining the human body anti-AQP4 autoantibodies detection material. According to the human body anti-AQP4 autoantibodies detection material and the preparation method thereof provided by the invention, the extracted AQP4-cell membrane complex is used as a detection material, so that the intrinsic protein amount causing background coloration can be reduced; compared with a method for using the cell as the detection material for carrying out immunofluorescence detection, the detection sensitivity and the specificity are further increased, a detection result is easier and clear to judge, and the AQP4 autoantibodies of a patient can be simply, conveniently and quickly detected.

Description

technical field [0001] The invention belongs to the field of biomedicine, and relates to a human body anti-AQP4 autoantibody detection material and a preparation method thereof. The prepared detection material can be used to detect human body anti-AQP4 autoantibody. Background technique [0002] Human autoantibodies are important serum markers of autoimmune diseases, and their rapid detection has important application value for the diagnosis, treatment and prognosis of autoimmune diseases. With the improvement of the sensitivity and specificity of autoantibody detection, the diagnostic criteria of related autoimmune diseases are also updated and adjusted accordingly. [0003] Neuromyelitis optica (neuromyelitis, NMO) is a serious demyelinating disease of the central nervous system, about 90% of NMO and more than half of patients with aquaporin 4 (aquaporin, AQP4) antibody positive. The discovery of aquaporin 4 antibody in 2004 provided objective evidence for NMO to become a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/12C12N15/85C07K14/47G01N33/531G01N33/543G01N33/68
CPCC07K14/47C12N15/85G01N33/531G01N33/54306G01N33/6893G01N2333/47G01N2800/28
Inventor 闫亚平柴单单李科
Owner SHAANXI MYBIOTECH CO LTD
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