Construction and application of humanized single-chain antibody libraries of cTnI (cardiac troponin I)

Abstract

The invention belongs to the field of gene engineering, and relates to a method for constructing mutant libraries of single-chain antibodies T of cTnI (cardiac troponin I). The method has the advantages that bacteria pRSF-Autodisplay are used as carriers, the capacity of each library can reach 1.5*10<5>, and the libraries are excellent in diversity as proved by DNA (deoxyribonucleic acid) sequencing; cTnI antigens from the fully humanized single-chain antibody libraries of the cTnI are used as targets by the aid of bacterium display technologies, and the humanized single-chain antibodies of the cTnI can be conveniently acquired after multiple screening cycles; the single-chain antibodies are recombinant antibodies, antibody heavy-chain variable regions and light-chain variable regions are connected with one another by a linker peptide by the aid of a gene engineering process to form each recombinant antibody, the single-chain antibodies are the smallest functional antibody fragments for keeping the antigen affinity activity and the specificity of parent antibodies, can obtained by means of in-vitro expression by the aid of gene engineering technologies and can be economically manufactured in the bacteria on a large scale, accordingly, immune detection antibodies can be easily, conveniently and economically manufactured, the detection reagent expenses can be greatly reduced, and the humanized single-chain antibody libraries are high in sensitivity and specificity, easy and convenient to operate and suitable for screening large quantities of samples, and have extremely broad application prospects.

Description

technical field [0001] The invention belongs to genetic engineering, and relates to the construction and application of a human cTnI single-chain antibody library. Background technique [0002] The development of antibody technology has gone through three stages. The first generation is antiserum polyclonal antibody. The second generation is the hybridoma monoclonal antibody (McAb) prepared by German scholar Kohler and British scholar Milstein using B lymphocyte hybridoma technology in the mid-1970s. This discovery is one of the milestones in the development of biotechnology, which has shown great importance in diagnosis, treatment, prevention and protein purification. However, because foreign proteins are easy to cause human anti-mouse antibody (HAMA) reaction, and human hybridoma technology is difficult to break through, the third generation of antibodies, genetically engineered antibodies, has emerged. Genetically engineered antibodies include three types, namely, chimer...

AI Technical Summary

Problems solved by technology

However, limited by the low transformation rate of yeast cells, it is necessary to establish multiple small libraries when constructing a yeast library, and then combine multiple small libraries, making the library construction process cumbersome and complicated.

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