Use of “one-pot, mix-and-read” peptide-MHC class I tetramers and predictive algorithms to improve detection of cytotoxic T lymphocyte responses in cattle

• Published in: Veterinary research (Paris) Vol. 45; no. 1; p. 50
• Main Authors: Svitek, Nicholas, Hansen, Andreas Martin, Steinaa, Lucilla, Saya, Rosemary, Awino, Elias, Nielsen, Morten, Buus, Søren, Nene, Vishvanath
• Format: Journal Article
• Language: English
• Published: London Springer-Verlag 28.04.2014; BioMed Central; BioMed Central Ltd
• Subjects: Algorithms; Amino Acid Sequence; Animals; beta 2-Microglobulin; beta 2-Microglobulin - immunology; Cattle; cattle diseases; cytotoxic T-lymphocytes; enzyme-linked immunosorbent assay; Enzyme-Linked Immunosorbent Assay - veterinary; epitopes; Epitopes, T-Lymphocyte; Epitopes, T-Lymphocyte - immunology; Histocompatibility Antigens; Immunology; Life Sciences; Major Histocompatibility Complex; Medical research; Medicine; Medicine & Public Health; Medicine, Experimental; Microbiology; Molecular Sequence Data; parasites; pathogens; Physiological aspects; prediction; Sequence Alignment; Sequence Alignment - veterinary; T-Lymphocytes, Cytotoxic; T-Lymphocytes, Cytotoxic - immunology; Theileria parva; Theileria parva - immunology; theileriosis; veterinary; Veterinary Medicine/Veterinary Science; Virology; Peptide Binding Specificity; Major Histocompatibility Complex Class; Major Histocompatibility Complex; Major Histocompatibility Complex Molecule; East Coast Fever
• ISSN: 1297-9716; 0928-4249; 1297-9716
• DOI: 10.1186/1297-9716-45-50

Peptide-major histocompatibility complex (p-MHC) class I tetramer complexes have facilitated the early detection and functional characterisation of epitope specific CD8⁺ cytotoxic T lymphocytes (CTL). Here, we report on the generation of seven recombinant bovine leukocyte antigens (BoLA) and recombinant bovine β2-microglobulin from which p-MHC class I tetramers can be derived in ~48 h. We validated a set of p-MHC class I tetramers against a panel of CTL lines specific to seven epitopes on five different antigens of Theileria parva, a protozoan pathogen causing the lethal bovine disease East Coast fever. One of the p-MHC class I tetramers was tested in ex vivo assays and we detected T. parva specific CTL in peripheral blood of cattle at day 15-17 post-immunization with a live parasite vaccine. The algorithm NetMHCpan predicted alternative epitope sequences for some of the T. parva CTL epitopes. Using an ELISA assay to measure peptide-BoLA monomer formation and p-MHC class I tetramers of new specificity, we demonstrate that a predicted alternative epitope Tp2₂₉₋₃₇ rather than the previously reported Tp2₂₇₋₃₇ epitope is the correct Tp2 epitope presented by BoLA-6*04101. We also verified the prediction by NetMHCpan that the Tp5₈₇₋₉₅ epitope reported as BoLA-T5 restricted can also be presented by BoLA-1*02301, a molecule similar in sequence to BoLA-T5. In addition, Tp5₈₇₋₉₅ specific bovine CTL were simultaneously stained by Tp5-BoLA-1*02301 and Tp5-BoLA-T5 tetramers suggesting that one T cell receptor can bind to two different BoLA MHC class I molecules presenting the Tp5₈₇₋₉₅ epitope and that these BoLA molecules fall into a single functional supertype.