AbstractSpontaneous cytolytic activity of shark peripheral blood leukocytes is observed only during periods of decreased environmental temperature (<23°C). The effector cell is adherent to glass and is phagocytic. Leukocytes tested during warmer periods (26–31°C) exhibit no spontaneous activity; however, glass‐adherent cells isolated from those fish are cytotoxicin vitro, indicating that the effector cell is present at all temperatures. During warmer temperatures, nonadherent cells added to adherent cells were shown to inhibit spontaneous cytotoxicity. This inhibition requires viable cells in contact with the spontaneous cytotoxic population. Thus decreased environmental temperature correlated with spontaneous cytotoxicity, and appears to affect a regulatory cell that is glass nonadherent. In addition, the cytotoxic effector cell is more active at 23°C than 30°Cin vitro. These data show that by the time of emergence of the nurse shark, a temperature‐dependent mechanism had evolved for cellular regulation of at least one immune function, spontaneous cyt