Preparation method of completely humanized antibody of infectious disease pathogen

Abstract

The invention relates to a preparation method of the completely humanized antibody of a infectious disease pathogen, which mainly comprises the following steps of: preparing an infectious disease pathogen antigen; fluorescently labeling an infectious disease pathogen antibody; concentrating and purifying the peripheral B cells of an infectious disease patient; combining the labeled pathogen antigen and the antigenic-specificity B cells; screening antigenic-specificity single B cells through a flow cytometer; carrying out single cell RT-PCR (Reverse Transcription-Polymerase Chain Reaction) amplification on genes positioned in the heavy chain and light chain variable region of the infectious disease pathogen antibody; connecting with a heavy chain and light chain constant region to construct the eukaryotic expression vector of the humanized antibody; and carrying out the high-efficiency expression and purification of a recombined antibody in an eukaryotic cell. The humanized monoclonal antibody prepared through the method disclosed by the invention can be used for the infectious disease diagnosis and the infectious disease treatment.

Description

technical field [0001] The invention relates to the technical field of antibody preparation, in particular to a method for preparing fully humanized antibodies to infectious disease pathogens. Background technique [0002] The development of monoclonal antibody drugs has gone through four stages: murine monoclonal antibody, chimeric monoclonal antibody, humanized monoclonal antibody and fully humanized monoclonal antibody. In 1975, German scholar Kohler and British scholar Milstein fused mouse myeloma cells and mouse spleen cells immunized with sheep red blood cells in vitro to form hybridoma cells, which inherited the genes of the two parental cells. It has both the characteristics of unlimited reproduction of myeloma cells and the ability to synthesize and secrete antibodies. Using this cell population derived from the proliferation of a single fused cell, specific monoclonal antibodies directed against an antigenic determinant can be prepared. There are some problems ...

AI Technical Summary

Problems solved by technology

The current phage antibody library technology also has shortcomings, such as the low affinity of antibodies obtained from the antibody library of unimmunized animals, the limitation of the conversion rate of exogenous genes, and the limited capacity of the antibody library to cover the antibody diversity of some animals, etc. ; and for ribosome display technology, how to further improve the stability of the system, especially how to prevent the degradation of mRNA and form a stable protein-ribosome-mRNA trimer is undoubtedly the key issue of the technology, how to improve The display of macromolecular proteins on ribosomes is also an issue that needs attention in future research