Cinemicroscopic studies of lymphocyte behavior in semi-solid medium: the polyclonal origin of lymphocyte colonies

Time-lapse cinemicroscopy was used to observe the development of lymphocyte colonies in a phytohemagglutinin-dependent one-step, two-layer agar culture system. More than 1000 h of culture time were recorded in a total of 14 independent experiments. Blast formation and organization of lymphocytes into highly motile pairs and clusters occurred early in culture (0-3 days). An increase in thymidine uptake also preceded the first detectable proliferation by 24 h. Mature lymphocyte colonies were found to be dynamic entities characterized by the continuous influx and egress of highly motile cells. Cell clusters and entire colonies were observed to locomote and on several occasions fuse to form larger structures. Macrophagelike cells located centrally within colonies appeared to play a major role in such behavior. Taken together these results conclusively demonstrate that, in the present system, lymphocyte colonies are the product of a complex pattern of cell interaction, proliferation, and cell motility and, as such, are polyclonal in origin.