Rapid identification of CD4+ T-cell epitopes using yeast displaying pathogen-derived peptide library.
Identification of CD4+ T-cell epitopes is a critical step in studying and modulating the immune responses to tumors, infectious agents, and autoantigens. Here we report a facile, accurate, and high-throughput method for CD4+ T-cell epitope identification using yeast displaying pathogen-derived peptide library. A library of DNA fragments that encode all the possible peptides with 10-20 amino acids from the antigens (single antigenic proteins or pathogenic organisms) are fused to the gene encoding the restriction single-chain MHC class II molecule in a yeast display vector. The resultant library of recombinant yeast cells are analyzed by FACS to identify those containing peptides with high affinity towards the restriction MHC molecule, which are subsequently screened for their ability to induce antigen-specific T-cell activation. DNA sequence analysis of selected positive clones results in direct identification of the antigenic peptides. We show that this method can be used to rapidly pinpoint the HA(306-322) epitope from the haemagglutinin protein and the entire influenza virus X31/A/Aichi/68 genome, respectively.
Wen
Fei
F
Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801, United States.
Esteban
Olga
O
Zhao
Huimin
H
D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D003001:Cloning, Molecular; D018604:Epitope Mapping / Q000379:methods*; D018984:Epitopes, T-Lymphocyte / Q000032:analysis* / Q000302:isolation & purification; D005434:Flow Cytometry; D015798:HLA-DR1 Antigen / Q000276:immunology*; D006388:Hemagglutinins / Q000276:immunology; D006801:Humans; D009980:Influenza A virus / Q000276:immunology; D008213:Lymphocyte Activation; D019151:Peptide Library; D010957:Plasmids / Q000235:genetics / Q000276:immunology; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000276:immunology
D018984:Epitopes, T-Lymphocyte; D015798:HLA-DR1 Antigen; D006388:Hemagglutinins; D019151:Peptide Library