Construction method of fully human phage ScFv natural antibody library and application thereof

Abstract

The invention relates to the technical field of biology, and discloses a construction method of phage ScFv antibody library and an application thereof. The phage ScFv antibody library construction method comprises the steps that 1, taking cDNA as a template, and amplifying a heavy chain variable region of an ScFv antibody and a light chain variable region of an induced ScFv antibody through PCR; 2) taking PCR products of a heavy chain variable region and a light chain variable region of the ScFv antibody as templates, and amplifying an ScFv antibody sequence by utilizing a primer with an SfiIrestriction enzyme cutting site through overlap PCR; and 3) performing double enzyme digestion on the pSEX and overlap PCR products of the vector by using SfiI restriction endonuclease, connecting tothe corresponding position of the pSEX vector after enzyme digestion, recovering the connection product, and performing electric conversion to obtain the phage ScFv antibody library. The phage ScFv antibody library obtained by the construction method provided by the invention has the characteristics of large library capacity, rich diversity and the like, and can be used for screening various specific antigens and specific humanized monoclonal antibodies of autoimmune disease related pathogens.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for constructing a fully human phage ScFv natural antibody library and its application. Background technique [0002] Antibody molecules are the most widely used protein molecules in the fields of biology and medicine. According to the survey report of the American Pharmaceutical Industry Association, the research and development ratio of monoclonal antibody drugs ranks first among all medical biotechnology products. As a reusable therapeutic drug, due to the immunogenicity of mouse monoclonal antibody, as a therapeutic product, fully human antibody has become the main direction of antibody development. There are many ways to produce human antibodies, such as the immortalization of B cells, hybridoma technology, and transgenic animals are all technical methods for producing human antibodies. Among them, there is a large amount of screening for the way to obtain antibodies us...

AI Technical Summary

Problems solved by technology

There are many ways to produce human antibodies, such as the immortalization of B cells, hybridoma technology, and transgenic animals are all technical methods for producing human antibodies. Among them, there is a large amount of screening for the way to obtain antibodies using traditional hybridoma technology, especially in the treatment There are human anti-mouse antibody reactions, which can reduce the curative effect and even cause allergic reactions, and these techniques are complicated to operate, difficult and expensive, and antibody library technology has become a popular method for preparing therapeutic drugs due to its advantages of simplicity, economy and speed. An important pathway for antibodies

The non-immune antibody libraries that have been reported so far mainly include natural antibody libraries, semi-synthetic antibody libraries and fully synthetic antibody libraries. During the construction of semi-synthetic and fully synthetic antibody libraries, antibody genes involve artificial synthesis, which requires computer modeling. Large, and may produce some invalid clones, affecting the quality of the antibody library

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