Experiments were done in vitro with cells derived from a C3H mouse mammary adenocarcinoma (MADCAP-37) to determine the influence of pH and sequencing on hyperthermia induced radiosensitization. The heat treatment was one hour at 42.5 degrees C in a precision controlled water bath. The heat treatment did not significantly reduce cell survival. Five Gy of x radiation was given at variable intervals from two hours before to two hours following heating. Survival curves under acid pH (6.4-6.7) and alkaline pH (7.2-7.4) were obtained for selected intervals, corresponding to points of interest in the sequencing study. Survival was greater for irradiation alone at acid pH than at alkaline pH. The greatest cell killing occurred when irradiation and hyperthermia were given simultaneously at either alkaline or acid pH. At alkaline pH, survival increased as the interval increased between heating and irradiation. At acid pH, radiation survival remained low for as much as two hours following heating. If tumors are at acid pH, the greatest differential effect should be obtained for irradiation following heating because cells at alkaline pH (normal) had approximately 10 times greater survival. Vascular changes caused by heating might alter that response.