Method for screening and identifying epitope of hepatitis b virus specific cytotoxic T lymphocyte

Abstract

The invention discloses a method for screening and identifying epitopes of hepatitis b virus specific cytotoxic T lymphocytes, which comprises the following steps: 1) preparing an artificial antigen presenting cell (aAPC) which loads any polypeptide; 2) loading the aAPC with epitope peptides; 3) allowing the aAPC loaded with the antigenic peptides to induce HBV antigen-specific CTL in vitro; 4) studying the anti-HBV function of the HBV antigen-specific CTL obtained in step 3) with a cell expressing HBV antigen as a target cell, wherein a cell expressing HBV antigen is used as a target cell which is a HepG2.2.15 cell; performing CTL killing experiment, HBV surface antigen determination, and HBV related gene DNA determination, respectively. The method is applicable to large-scale screening and identification of CTL epitopes.

Description

technical field [0001] The invention belongs to the fields of cell biology and immunology of biotechnology, and relates to using cells expressing hepatitis B virus (Hepatitis B virus, HBV) protein as target cells, and coating artificial antigen-presenting cells with candidate epitope peptides ( Artificial antigen-presenting cells (aAPC) induce HBV-specific cytotoxic T lymphocytes (Cytotoxic T Lymphocytes, CTL) in vitro as effector cells, and observe the effect of effector cells on killing target cells to screen and identify HBV-specific CTL epitopes. Background technique [0002] The treatment of chronic hepatitis B virus infection is still a major problem in the medical field. The current anti-HBV treatment drugs cannot completely eliminate the virus. How to effectively eliminate HBV is still a hot spot in the treatment of hepatitis B. [0003] (1) HBV-specific CTL is an important factor for the body to clear HBV. It is of great significance for the treatment of chronic he...

AI Technical Summary

Problems solved by technology

[0007] However, there are still some problems in the study of HBV-specific CTL epitopes: ①, synthetic peptides covering the entire sequence of the HBV antigen system are not used, and some important information may be missed; ②, only high affinity with HLA (human leukocyte antigen) molecules is selected The analysis of peptides that bind sex may miss peptides with low affinity but strong immunogenicity; ③ Most studies only use limited commonly used epitopes to immunize CTLs in patients with different clinical types of hepatitis B At present, there are a large number of mutant viruses appearing clinically, and the sequences of these epitopes also change, so these epitopes cannot be used in the research of many patients infected with mutant viruses; ④, there is almost no HLA-restricted HBV CTL The epitope was reported by my country, and the HBV genotype of my country is very different from that of European and American countries

[0008] Problems in the methodology of epitope research: In the past, cloning technology was used to determine the immune epitope by gradually narrowing the screening range, and animal (or human) experiments were used to identify it, which was time-consuming, labor-intensive, and inefficient

With the deepening of the understanding of HLA, the epitope that may exist in a certain antigen can be predicted by bioinformatics according to the epitope binding motif of different HLA molecules, but it must be identified in combination with modern experimental techniques