BLASTOGENESIS AND INTERLEUKIN-2 RECEPTOR EXPRESSION ASSAYS IN THE HARBOR SEAL (PHOCA-VITULINA)

Two in vitro functional assays were developed to evaluate mitogen-indu ced responses of peripheral blood mononuclear leukocytes (PBML) from f ree-ranging harbor seals, Phoca vitulina. Lymphocyte proliferation was measured by a standard blastogenesis assay following optimization of culture conditions including mitogen concentration, cell density, and incubation time. These optimized parameters, with the exception of inc ubation time, were subsequently employed to measure lymphocyte activat ion by analytical now cytometry using fluorochrome-based identificatio n of cell surface interleukin-2 receptor (IL-2r) expression. Baseline values established for free-ranging harbor seals had extensive animal variability; there was evidence that the samples were derived from a g roup of animals with a normal distribution. Positive correlations were observed between blastogenesis assays, and between blastogenesis and activation assays, when using pokeweed or concanavalin A as the stimul us. However, no relationship was found in the expression of the IL-2r induced by these mitogens. This result supports the contention that th e two mitogens stimulate different lymphocyte subpopulations. This was observed only with the IL-2r expression assay because of its unique a bility to measure the number of T lymphocytes initially activated rath er than the ultimate number of progeny cells identified by blastogenes is. Both assays, used concurrently, should provide a more comprehensiv e representation of lymphocyte competence and serve as a measure of an imal health.