Methods devised for generation and assay of migration inhibitory factor (MIF) on a reduced scale have permitted the performance of experiments which demonstrate that glass bead column-purified guinea pig lymph node lymphocytes do not produce MIF or proliferate in response to antigen. Evidence that the macrophage is the essential cellular element eliminated by the purification procedure is demonstrated by the restoration of both lymphocyte responses upon addition of 5% macrophages to the purified lymphocytes. Autologous, immune and syngeneic, non-immune peritoneal exudate macrophages were equally effective in restoring MF production by purified lymphocytes. Supernatants which inhibited the migration of peritoneal exudate macrophages had only minimal or no effect on the migration of alveolar macrophages, confirming that the inhibitory effects studied were attributable to MF and not cytophilic antibody.