A scalable multiplex assay enabling assessment of T cell receptor specificity to hundreds of self- and pathogen-derived antigens

• Published in: The Journal of immunology (1950) 2016-05, Vol.196 (1_Supplement), p.209-209.4
• Main Authors: Klinger, Mark, Taniguchi, Ruth, Hu, Joyce, Hayes, Tim, Wittkop, Tobias, Asbury, Thomas, Moorhead, Martin, Emerson, Ryan, Sherwood, Anna, Robins, Harlan, Faham, Malek
• Format: Article
• Language: English
• ISSN: 0022-1767; 1550-6606
• DOI: 10.4049/jimmunol.196.Supp.209.4

Monitoring antigen-specific T cells is critical for the study of immune responses and development of biomarkers and immunotherapeutics. We previously developed and validated a novel multiplex assay (MIRA, or Multiplexed Identification of T cell Receptor Antigen specificity) that combines conventional immune monitoring techniques and TCR repertoire sequencing to assess T cell specificity to large numbers of query antigens. MIRA is a sensitive assay enabling detection of antigen-specific TCR clonotypes well below the limit of detection of conventional immune monitoring assays including flow cytometry and ELISPOT. Here we report the results from a scaled-up version of the assay using 270 different query peptide antigens (159 self- and 111 pathogen-derived). We identified >500 TCR clonotypes at frequencies as low as 1 per million T cells that were specific to 41 query antigens from 6 healthy HLA-A*02-positive individuals. Most of the antigen-specific TCRs identified recognized one of 27 different peptides derived from a variety of pathogens including CMV, EBV, Flu, Rotavirus, HSV, mTB, WNV and HIV. A subset of antigen-specific TCRs recognized one of 14 different peptides derived from self including MART1, RCC, BCL-2, MAGE, STEAP1, KLK4, CAMEL and MOG. These data support the notion that escape and survival of self antigen-specific T cells occurs without causing overt autoimmunity in healthy individuals. We show here that MIRA can be used to assess TCR specificities to hundreds of query antigens simultaneously. The assay is highly scalable and can be easily modified to accommodate thousands of additional query antigens. This technology may be used to monitor T cell specificity to antigens relevant to infection, autoimmunity and cancer.