Peritoneal exudate lymphocytes (PEL) from immune guinea pigs adhere to macrophages carrying the relevant antigen and are thereby stimulated to proliferate in culture. The resultant PEL represent a population highly enriched with regard to their capacity to specifically rebind to antigen-pulsed macrophages. We have studied the mechanisms underlying specific binding of lymphocytes to macrophages by examining the effects of physical and chemical modifications of the two cell types. Specific binding was inhibited by fixation of cells, metabolic inhibitors, low temperatures, cytochalasin B and divalent cation depletion. After specific binding has taken place, cation depletion, but not cytochalasin B or low temperatures, disrupts binding. These observations indicate that specific binding occurs by a series of discrete events that can be operationally distinguished.