title: Parallel detection of antigen-specific T cell responses by combinatorial encoding of MHC multimers.
abstract: Fluorescently labeled multimeric complexes of peptide-MHC, the molecular entities recognized by the T cell receptor, have become essential reagents for detection of antigen-specific CD8(+) T cells by flow cytometry. Here we present a method for high-throughput parallel detection of antigen-specific T cells by combinatorial encoding of MHC multimers. Peptide-MHC complexes are produced by UV-mediated MHC peptide exchange and multimerized in the form of streptavidin-fluorochrome conjugates. Eight different fluorochromes are used for the generation of MHC multimers and, by a two-dimensional combinatorial matrix, these eight fluorochromes are combined to generate 28 unique two-color codes. By the use of combinatorial encoding, a large number of different T cell populations can be detected in a single sample. The method can be used for T cell epitope mapping, and also for the monitoring of CD8(+) immune responses during cancer and infectious disease or after immunotherapy. One panel of 28 combinatorially encoded MHC multimers can be prepared in 4 h. Staining and detection takes a further 3 h.
authors.affiliation: Center for Cancer Immune Therapy, Department of Hematology, University Hospital Herlev, Denmark.
authors.forename: Rikke Sick
authors.initials: RS
authors.lastname: Andersen
authors.forename: Pia
authors.initials: P
authors.lastname: Kvistborg
authors.forename: Thomas Mørch
authors.initials: TM
authors.lastname: Frøsig
authors.forename: Natasja W
authors.initials: NW
authors.lastname: Pedersen
authors.forename: Rikke
authors.initials: R
authors.lastname: Lyngaa
authors.forename: Arnold H
authors.initials: AH
authors.lastname: Bakker
authors.forename: Chengyi Jenny
authors.initials: CJ
authors.lastname: Shu
authors.forename: Per thor
authors.initials: Pt
authors.lastname: Straten
authors.forename: Ton N
authors.initials: TN
authors.lastname: Schumacher
authors.forename: Sine Reker
authors.initials: SR
authors.lastname: Hadrup
mesh_terms: D000818:Animals; D000941:Antigens / Q000737:chemistry; D018414:CD8-Positive T-Lymphocytes / Q000737:chemistry / Q000276:immunology*; D018604:Epitope Mapping / Q000379:methods*; D018984:Epitopes, T-Lymphocyte / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000032:analysis; D006183:H-2 Antigens / Q000378:metabolism*; D006680:HLA Antigens / Q000378:metabolism*; D006801:Humans; D051379:Mice; D061848:Optical Imaging / Q000379:methods; D045663:Quantum Dots
chemical_list: D000941:Antigens; D018984:Epitopes, T-Lymphocyte; D005456:Fluorescent Dyes; D006183:H-2 Antigens; D006680:HLA Antigens