In vitro stimulation of bovine peripheral blood lymphocytes: standardization and kinetics of the response

A microculture system is described to study the in vitro properties of bovine peripheral blood lymphocytes. The use of a semiautomatic multiple sample harvester (SAMSH) enabled rapid multiple tests, using relatively few cells. Careful analysis of lymphocyte transformation to phytohemagglutinin (PHA) and pokeweed mitogen (PWM) determined optimal conditions as (a) 72 hour culture period, (b) 2.0 x 10⁵ cells per culture in RPMI 1640 medium containing 15% heat inactivated autologous serum, and (c) 1 μCi of [³H] thymidine per culture added to cultures 18 hours before termination. The whole blood culture technique was also investigated and evaluated. Normal responses of cattle are reported.