Mitogenig Responses of Sheep Peripheral Blood T-Lymphocytes To Pha and Lps in a Hanging Drop Microassay

Culture conditions for studying sheep PBL responses to mitogens (PHA and LPS) in a hanging drop micro-assay were investigated. Using those conditions, the responses of putative sheep T-cells (vide infra) to the two mitogens were examined.The results show that sheep PBL at a cell density of 1–2×10⁶/ml responded optimally to LPS (50 (µg/ml) and PHA (3.125 µg/ml) when cultured in RPMI-1640 in the presence of 2ME, 5% FGS for 48 to 146 h. The cells were cultured as hanging drops (20 µl) in Terasaki plates. Lamb serum failed to support responses to LPS.T-cells prepared by the nylon wool technique and Helix pomatia-Sepharose-6MB column chromatography responsed vigorously to PHA, but responses to LPS were weak.