Infection of T. annulata -transformed cells with BHV-1, recombinant vaccinia virus NYVAC-gD, and recombinant Tp1-canarypox virus. (a). T. annulata transformed cells from animal 03430 were harvested 18 h after infection with BHV-1 or with NYVAC-gD and stained by indirect immunofluorescence with a monoclonal antibody specific for the BHV-1 protein gD. Control uninfected cells stained with the same antibody are shown in the left panel. Infected cells stained only with secondary antibody were also negative (data not shown). The percentages of positive cells are indicated within the panels. (b). Cells infected with Tp1-canarypox were tested for recognition by a CD8 T cell line specific for Tp1 from a T. parva- immune, class I MHC A18-homozygous cow, using an IFN-γ release assay. Results are shown for T cells stimulated with autologous or allogeneic T. parva transformed cells (Tp and Tp allo respectively), or autologous T. annulata-transformed cells pulsed with Tp1 214-224 peptide (Ta-Tp1 P), or infected with a canarypox virus expressing Tp1 (Ta-Tp1V), canarypox virus expressing Tp2 (Ta-Tp2 V) or uninfected (TA). Supernatants of T cell cultures collected 20 h after antigenic stimulation were assayed for IFN-γ using a specific ELISA. The results represent the mean of measurements from duplicate cultures. The experiment was repeated once with similar results.