Downloaded from orbit.dtu.dk on: Dec 14, 2021 CD4+ T-cell lines used to evaluate a Mycobacterium avium subsp. paratuberculosis (MAP) peptide vaccine Lybeck, Kari; Sjurseth, Siri K. ; Al-Touama, Zainab; Melvang, Heidi Mikkelsen; Aagaard, Claus; Lundegaard, Claus; Jungersen, Gregers; Andersen, Peter; Olsen, Ingrid; Tollefsen, Stig Publication date: 2015 Document Version Publisher's PDF, also known as Version of record Link back to DTU Orbit Citation (APA): Lybeck, K., Sjurseth, S. K., Al-Touama, Z., Melvang, H. M., Aagaard, C., Lundegaard, C., Jungersen, G., Andersen, P., Olsen, I., & Tollefsen, S. (2015). CD4+ T-cell lines used to evaluate a Mycobacterium avium subsp. paratuberculosis (MAP) peptide vaccine. Abstract from 5th European Veterinary Immunology Workshop, Vienna, Austria. General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights.  Users may download and print one copy of any publication from the public portal for the purpose of private study or research.  You may not further distribute the material or use it for any profit-making activity or commercial gain  You may freely distribute the URL identifying the publication in the public portal If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim. CD4+ T-cell lines used to evaluate a Mycobacterium avium subsp. paratuberculosis (MAP) peptide vaccine Kari R. Lybeck1, Siri K. Sjurseth1, Zainab Al-Touama1, Heidi Mikkelsen2, Claus Aagaard3, Claus Lundegaard2, Gregers Jungersen2, Peter Andersen3, Ingrid Olsen1, Stig Tollefsen1. 1 Section for Immunology, Norwegian Veterinary Institute, Oslo, Norway. Section for Immunology and Vaccinology ,Technical University of Denmark, Copenhagen, Denmark. 3 Vaccine Research & Development, Statens Serum Institut, Copenhagen, Denmark. 2 The aim of the study was to establish a protocol for generation of MAP-specific T-cell lines and to use these lines for evaluation of a peptide vaccine. A protocol for culturing T-cell lines from peripheral blood of goats naturally infected with MAP was established. CD4+ T cells were positively selected using an anti CD4 mAb and Dynabeads. Sorted CD4+ cells were cultivated with purified protein derivative from MAP (PPDj) or E. coli sonicate, IL-2, and IL-15. After two cultivation cycles, T cells were tested for recall responses in a proliferative T-cell assay. T-cell line responses were in average 92 % for PPDj, and -3 % for E. coli sonicate. CD4+ T-cell lines stimulated with PPDj showed a 6 fold increase in IFN- γ production compared to controls. These results indicated that the T-cell lines were MAP-specific. The protocol was subsequently used to evaluate MAP-specific peptides as vaccine antigens. T-cell lines were now generated by cultivating CD4+ cells with peptides instead of PPDj. Initially, both healthy and MAP-infected goats were vaccinated with 119 peptides defined by in silico analysis. Cellular responses to the peptides were not detected using standard IFN- γ plasma ELISA. However, testing of T-cell lines from the MAP-infected goats identified peptides that induced strong proliferative responses. The 23 peptides inducing the strongest responses were used in a second vaccination trail with healthy goat kids. Vaccinated kids developed strong IFN-γ and antibody responses, and these MAP-specific peptides show great potential for use in a subunit vaccine. Generation of T-cell lines was a valuable tool for selecting MAP vaccine antigens, and the protocol can also be applied for identifying vaccine candidates for other diseases.