Direct phenotypic analysis of human MHC class I antigen presentation: visualization, quantitation, and in situ detection of human viral epitopes using peptide-specific, MHC-restricted human recombinant antibodies.
The advent in recent years of the application of tetrameric arrays of class I peptide-MHC complexes now enables us to detect and study rare populations of Ag-specific CD8(+) T cells. However, available methods cannot visualize or determine the number and distribution of these TCR ligands on individual cells nor detect APCs in tissues. In this study, we describe for the first time studies of human class I peptide-MHC ligand presentation. These studies were facilitated by applying novel tools in the form of peptide-specific, HLA-A2-restricted human recombinant Abs directed toward a viral epitope derived from human T cell lymphotropic virus type I. Using a large human Ab phage display library, we isolated a large panel of recombinant Fab Abs that are specific for a particular peptide-MHC class I complex in a peptide-dependent, MHC-restricted manner. We used these Abs to visualize the specific complex on APCs and virus-infected cells by flow cytometry, to quantify the number of, and visualize in situ, a particular complex on the surface of APCs bearing complexes formed by naturally occurring active intracellular processing of the cognate viral Ag. These findings demonstrate our ability to transform the unique fine specificity, but low intrinsic affinity of TCRs into high affinity soluble Ab molecules endowed with a TCR-like specificity toward human viral epitopes. These molecules may prove to be crucial useful tools for studying MHC class I Ag presentation in health and disease as well as for therapeutic purposes in cancer, infectious diseases, and autoimmune disorders.
Cohen
Cyril J
CJ
Faculty of Biology, Technion-Israel Institute of Technology, Haifa, Israel.
Sarig
Offra
O
Yamano
Yoshihisa
Y
Tomaru
Utano
U
Jacobson
Steven
S
Reiter
Yoram
Y
D000595:Amino Acid Sequence; D000915:Antibody Affinity; D000918:Antibody Specificity; D017951:Antigen Presentation / Q000276:immunology*; D000938:Antigen-Presenting Cells / Q000276:immunology / Q000378:metabolism; D001666:Binding Sites, Antibody; D001667:Binding, Competitive / Q000276:immunology; D002460:Cell Line; D002461:Cell Line, Transformed; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D018984:Epitopes, T-Lymphocyte / Q000276:immunology* / Q000378:metabolism; D016356:Gene Products, tax / Q000276:immunology* / Q000378:metabolism; D015789:HLA-A2 Antigen / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D015368:Human T-lymphotropic virus 1 / Q000276:immunology*; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000378:metabolism*; D016130:Immunophenotyping / Q000379:methods*; D007424:Intracellular Fluid / Q000276:immunology / Q000378:metabolism; D008024:Ligands; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000276:immunology* / Q000378:metabolism; D011948:Receptors, Antigen, T-Cell / Q000276:immunology / Q000378:metabolism; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism; D016176:T-Lymphocyte Subsets / Q000276:immunology / Q000378:metabolism / Q000821:virology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology / Q000378:metabolism / Q000821:virology; D014407:Tumor Cells, Cultured
D018984:Epitopes, T-Lymphocyte; D016356:Gene Products, tax; D015789:HLA-A2 Antigen; D007140:Immunoglobulin Fab Fragments; D008024:Ligands; D010446:Peptide Fragments; D011948:Receptors, Antigen, T-Cell; D011994:Recombinant Proteins