TY - CHAP
T1 - Construction and screening of an antigen-derived peptide library displayed on yeast cell surface for CD4+ T cell epitope identification
AU - Wen, Fei
AU - Zhao, Huimin
PY - 2013
Y1 - 2013
N2 - Identification of T cell epitopes is a critical, but often difficult step in studying T cell function and developing peptide-based vaccines and immunotherapies. Unlike antibodies that recognize free soluble antigens, T cell receptor (TCR) recognizes its epitope bound to major histocompatibility complex (MHC) expressed on antigen presenting cells (APCs). In addition, the examination of T cell epitope activity requires the use of professional APCs, which are difficult to isolate, expand, and maintain. To address these issues, we have developed a facile, accurate, and high-throughput method for T cell epitope mapping by screening antigen-derived peptide libraries in complex with MHC protein displayed on yeast cell surface. Here, we use hemagglutinin and influenza A virus X31/A/Aichi/68 as examples to describe the key steps in identification of CD4+ T cell epitopes from a single antigenic protein and the entire genome of a pathogen, respectively. Methods for single-chain peptide-MHC complex vector design, yeast surface display, peptide library generation in Escherichia coli, and functional screening in Saccharomyces cerevisiae are discussed.
AB - Identification of T cell epitopes is a critical, but often difficult step in studying T cell function and developing peptide-based vaccines and immunotherapies. Unlike antibodies that recognize free soluble antigens, T cell receptor (TCR) recognizes its epitope bound to major histocompatibility complex (MHC) expressed on antigen presenting cells (APCs). In addition, the examination of T cell epitope activity requires the use of professional APCs, which are difficult to isolate, expand, and maintain. To address these issues, we have developed a facile, accurate, and high-throughput method for T cell epitope mapping by screening antigen-derived peptide libraries in complex with MHC protein displayed on yeast cell surface. Here, we use hemagglutinin and influenza A virus X31/A/Aichi/68 as examples to describe the key steps in identification of CD4+ T cell epitopes from a single antigenic protein and the entire genome of a pathogen, respectively. Methods for single-chain peptide-MHC complex vector design, yeast surface display, peptide library generation in Escherichia coli, and functional screening in Saccharomyces cerevisiae are discussed.
KW - CD4 T cell epitope mapping
KW - Flow cytometry
KW - High throughput screening
KW - HLA-DR1
KW - Influenza A virus
KW - Major histocompatibility complex
KW - Peptide library
KW - Single-chain peptide-MHC complex
KW - Yeast display
UR - http://www.scopus.com/inward/record.url?scp=84884182990&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84884182990&partnerID=8YFLogxK
U2 - 10.1007/978-1-62703-589-7_15
DO - 10.1007/978-1-62703-589-7_15
M3 - Chapter
C2 - 23963942
AN - SCOPUS:84884182990
SN - 9781627035880
T3 - Methods in Molecular Biology
SP - 245
EP - 264
BT - Immunoproteomics
A2 - Fulton, Kelly
A2 - Twine, Susan
ER -