The effects of incubation at mildly elevated temperatures on HIV-1 inactivation and in vitro red blood cell properties were investigated. Red cells (55% Hct) were leukodepleted (3 log(10)) by filtration, maintained at 45 or 47 °C for 4 or 8 h, and then stored at 4°C. Hemolysis was twice that of controls after 42-day storage for samples treated for 4h at 45°C, and five times larger for samples heated at 47°C. There was also a significant increase in the rate of potassium loss, an early decrease in ATP levels, and an initial drop in pH for samples treated at either temperature. Larger differences were observed for samples exposed to these elevated temperatures for 8 h. Osmotic deformability curves obtained by ektacytometry showed dramatic decreases in red cell deformability at both temperatures and for both time periods. HIV-1 inactivation in red cells treated at 45 °C (approximately 0.25 log(10)/h) was considerably less than that obtained in tissue culture medium (1-2 log(10)/h). Since the decrease in red cell deformability is likely to indicate reduced red cell function and survival, and the rate of HIV-1 inactivation is low, mild heat treatment is not an adequate process for viral inactivation of red cell products.