We studied the role of stimulant processing and presentation and of IL 1 in monocyte-mediated activation of human lymphoproliferative responses. The effects of two lysosomotropic agents, ammonium chloride and chloroquine, on the capacity of human monocytes to activate T lymphocyte responses to the soluble antigen streptolysin O (SLO) and to the polyclonal stimulant S. aureus protein A (SpA) were investigated. These agents inhibited the presentation of SLO and SpA by human monocytes in a dose-dependent manner. The inhibition occurred if monocytes were treated with ammonium chloride and chloroquine for 1.5 hr, starting only 30 min after exposure to the stimulants, whereas only minimal inhibition occurred when monocytes were treated with the two lysosomotropic compounds 2 hr after pulsing with SLO or SpA. In contrast, cell membrane alloantigen presentation by monocytes in the MLR was not affected by ammonium chloride or chloroquine treatment. Thus, these reversible inhibitors of monocyte phagosome-lysosome functions presumably interfere with intracellular processing of the stimulants but do not seem to interfere with alloantigen presentation at the cell surface. Furthermore, we investigated whether gently fixed monocytes were still capable of passively presenting stimulant or whether active metabolic processes as well as IL 1 were required. We observed that only monocytes treated with paraformaldehyde after SLO or SpA pulsing stimulated a proliferative response by T lymphocytes, provided 50 U/ml of partially purified human IL 1 were added back to cultures. In contrast, monocytes fixed before exposure to SLO or SpA were not able to stimulate T lymphocytes even if supplemented by IL 1. Taken together these data suggest that a finite incubation period is required for human monocytes to become able to present SLO or SpA to T lymphocytes. During this time the soluble stimulants presumably undergo some metabolic process in viable macrophages perhaps at the phagosome-lysosome level, to become recognizable by T lymphocytes.