Peritoneal exudate lymphocyte. III. Dissociation of antigen reactive lymphocytes from antigen binding cells in a T lymphocyte enriched population in the guinea pig

Guinea pig peritoneal exudate lymphocytes (PEL) are a population of highly antigen reactive thymus derived (T) lymphocytes that are relatively macrophage depleted (<1 to 2%). The authors attempted to determine if these T lymphocytes, as a consequence of their extreme antigen sensitivity, could directly bind sufficient amounts of antigen (PPD) to become activated and thereby exhibit macrophage independent proliferation in vitro. Using a functional assay of antigen binding they determined that the antigen binding ability of macrophages is so great that all of the antigen binding in the PEL population can be accounted for by the few contaminating macrophages present in the population. Repeated passage of the PEL population over nylon glass bead adherence columns results in a greater than 100 fold reduction in the number of residual macrophages. This degree of macrophage depletion is associated with a significant decrease in both the functional antigen binding ability (86%) and the antigen induced proliferative response (88%) of the highly purified PEL population. However, this loss of activity is due solely to the depletion of macrophages and not T lymphocytes, since the antigen induced proliferation of the purified population can be restored with fresh macrophages and antigen. The data demonstrate the extreme difficulty of completely removing macrophages from a complex lymphoid population such as the PEL and ascribing with certainty any remaining function solely to its component lymphocytes rather than its macrophages. These studies argue against binding of functional amounts of PPD directly by T lymphocytes and suggest that antigen binding by macrophages is an obligatory step in the activation of T lymphocytes in vitro.