From several studies of hyperthermia there have been reports that hypoxic cells are more sensitive to heat than their oxic counterparts. Experimental techniques in this investigation eliminate the effect of pH, trypsinization and cell attachment, when assaying the effects of hyperthermia on cells. Under hypoxic conditions, HeLa S3 and Chinese hamster cell-lines do not have an increased sensitivity to heat compared with oxic cells. HeLa S3 cells are protected against heat by hypoxia. Light-microscopy indicates the rupture of the plasma membrane, occasional nuclear budding, membrane vesicles and granulation of cell contents after heating at 43 degrees C for 3 hours. Scanning electron micrographs show that cells are more rounded after heat treatment and that there is an accompanying decrease in the number of microvilli, suggesting that the mechanism of cell attachment is affected. Heated cells should be delicately handled and subjected to the minimal trauma so that an accurate comparison of survival can be made.