Application of Multiparametric Flow Cytometry Panels to Study Lymphocyte Subpopulations in Tuberculin-Positive Cattle

Flow cytometry (FC) is widely used in microbiology, immunology, hematology, and oncology. In the veterinary field, FC enabled the study of the immune response in cattle infected with different pathogens, as well as vaccine testing. However, few fluorochrome-conjugated antibodies recognize bovine antigens, limiting the possible benefits of FC and the implementation of multiparametric analysis for more complex studies. Two cytometry panels with five colors each were designed and implemented for the study and identification of populations and subpopulations of T cells derived from the peripheral blood mononuclear cells of dairy heifers. Both panels detected differences in T cell subpopulations between heifers positively and negatively tested for tuberculin; they detected overexpression of CD25⁺ and CD45RO⁺ in tuberculin-positive heifers after stimulation with a culture filtrate protein extract (CFPE) from Mycobacterium bovis (M. bovis). We identified subpopulations of T cells from peripheral blood mononuclear cells using two multicolor panels. These panels could be used to analyze total bovine blood in immunopathogenic studies and vaccine development. The same strategy could be implemented in other species of veterinary interest.