Flow cytometric analysis of mitogen-induced activation of rainbow trout (Oncorhynchus mykiss) peripheral blood leucocytes

Proliferation of rainbow trout peripheral blood leucocytes in vitro is usua lly assessed by measuring incorporated tritiated thymidine. In this report we monitored the in vitro proliferative response to the mitogen Concanavali n A (Con A) by means of flow cytometry (FCM) and H-3-thymidine incorporatio n. When analysed by FCM, blood leucocytes displayed two main cell populatio ns with distinct forward and side scatter (FSC/SSC) characteristics: lympho cytes with low FSC/SSC values and non-lymphoid leucocytes (NLL) with increa sed FSC/SSC values. The nature of these cell types were confirmed by micros copy. Interestingly, the FSC/SSC pattern of lymphocytes remained unchanged after in vitro stimulation with Con A, whereas cells from the NLL populatio n showed a marked shift towards increased FSC values. In stimulated culture s, the increase of FSC values of the NLL population significantly correlate d with contemporarily measured H-3-thymidine incorporation (r = 0.7, P < 0. 001). The mitogenic response of blood leucocytes originating from different individual fish varied over wide ranges. It was found to be related to the numbers of NLL present in the leucocyte sample. The present results show t hat qualitative and quantitative FCM analysis of morphological parameters ( FSC/SSC) of blood leucocytes makes it possible to discriminate between leuc ocyte populations of the rainbow trout and to monitor cell proliferation ex periments.