Serum amyloid protein A (SAA) antigen epitope and predication and verification method thereof as well as preparation method of monoclonal antibody

Abstract

The invention relates to a predication and verification method of a serum amyloid protein A (SAA) antigen epitope and a preparation method of a monoclonal antibody. The predication and verification method comprises the following steps: predicating surface accessibility, an flexibility sequence region, a beta turn, antigenicity, hydrophilicity, a linear epitope and a long-fragment epitope of a serum amyloid protein A antigen to obtain a plurality of epitope sequences respectively; removing repeated epitopes in the plurality of epitope sequences and other sequences contained in other relativelylong epitopes to obtain a plurality of non-redundant predication epitopes; and verifying whether the plurality of non-redundant predication epitopes are identification epitopes of the antibody or notby adopting ELISA (Enzyme Linked Immunosorbent Assay). According to the invention, the problem that the predication accuracy of a single method is low is overcome; through experimental verification, the problem that the SAA antigen epitope is unknown is solved; and the monoclonal antibody is prepared by applying synthesized antigen epitope polypeptide, so that the problem that SAA is difficult topurify and obtain is overcome and the cost is reduced by one time or more.

Description

technical field [0001] The invention belongs to the field of preparation of in vitro diagnostic raw materials by biotechnology, belongs to the raw materials for the preparation of clinical diagnostic kits in the in vitro diagnostic industry, and specifically relates to a serum amyloid A antigen epitope, its prediction and verification method and a preparation method of a monoclonal antibody. Background technique [0002] Humoral immunity or antibody production is mainly mediated by B cells, and B cells recognize antigens through membrane-bound antibodies or B cell receptors (B cell receptors, BCRs). Editing of antibody genes and secretion of antibodies that bind to antigens to inactivate or eliminate them. [0003] Immunoinformatics is an informatics method that uses computer methods to apply and solve immune problems in immunology, and is a branch of bioinformatics. The two central goals of immunoinformatics are the prediction of B cell epitopes and T cell epitopes, which ...

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Problems solved by technology

[0009] 1. The current immunoinformatics methods and tools are mainly aimed at the general prediction of B cell epitopes from high-throughput data, and the prediction accuracy is low, and there is a lack of experimental screening and verification of antigen epitopes, and there is also a lack of application of validated antigen tables. Application of preparing antibodies in situ

[0010] 2. Up to now, there are no research reports on SAA epitopes, and the immunoinformatics analysis and follow-up experimental screening verification of SAA epitopes are also blank

[0011] 3. SAA is an emerging sensitive indicator that reflects the infection status of the body and the recovery of inflammation. However, the existing anti-SAA antibodies in the market are not high in sensitivity and specificity, and most of them are antibodies for scientific research. SAA antibodies for in vitro diagnosis can be applied Deficient, and the epitope recognized by the antibody determinant is unknown

[0012] 4. The half-life of SAA in human blood is about 1 day, and more than 95% of it is combined with lipids such as high-density lipoprotein and cholesterol. It also has interactions and interactions with various cell receptors, mucopolysaccharides, and cystatin C. In addition, free SAA in blood is easily degraded by protease, so it is very difficult to purify SAA in human blood

[0013] 5. Recombinant SAA (rSAA) is difficult to purify due to SAA's lipophilicity and is expensive, and recombinant SAA generally presents hexamers, tetramers, octamers, etc., lacks the natural conformation of human blood SAA, and has poor antigenicity

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