Changes in the structure of the surface of mastocytoma cells were induced by hyperthermia and were investigated by means of cell electrophoresis. A decrease in the cell electrophoretic mobility was detected as early as 15 min after treatment at 42 degrees and progressed more rapidly under hypoxic conditions than under oxic conditions. Subsequent recovery of electrophoretic mobility at 37 degrees was dependent on the length of heat treatment and oxygenation. The surviving fraction of cells detected by their colony-forming ability and the fraction of electrophoretically recovered cells 24 hr after exposure to hyperthermia showed a good statistical correlation. It was suggested that the mechanism of electrophoretic mobility reduction by heating was the vertical translocation of hyaluronidase-sensitive charge from the peripheral layer into a deeper layer by combined use of specific enzymes and stepwise different ionic strengths. These results suggest the importance of irreparable changes of membrane conformation in the loss of colony-forming ability of heated tumor cells.