Evaluation of cell replication by bovine T cells in polyclonally activated cultures using carboxyfluorescein succinimidyl ester ([formula omitted]) loading and flow cytometric analysis

Studies reported here demonstrated that carboxyfluorescein succinimidyl ester (CFSE) loading of lymphocytes and flow cytometric analysis is a powerful assay to assess the kinetics and extent of cellular replication by bovine T-cell subpopulations in heterogeneous cultures of peripheral blood mononuclear cells (PBMC) where subpopulation interactions can occur. As CFSE analysis allows determination of the proportion of lymphocytes that divided, as well as the number of cell divisions each cell underwent, distinctions in responses among mitogen-stimulated cultures could be made even when3H-thymidine incorporation was equivalent. When combined with surface staining for detection of differentiation antigens, differences among T-cell subpopulations with regard to the number of divisions their members had undergone, were found. Anti-CD3 mAb stimulated both CD8+and CD4+T cells to undergo several cell divisions in 72 hours, while there was essentially no division by γδ T cells. In contrast, in concanavalin A-stimulated cultures, all T-cell subpopulations had divided.