The sensitivity of different cell lines (Chinese hamster ovary, HeLa, L1210, and P388) to 43 degrees was compared Chinese hamster ovary and HeLa cells were much less temperature sensitive than were L1210 or P388 cells. This difference persisted even when HeLa and L1210 cells were grown in the same medium and suggested that this was an inherent difference between the cell lines. Of all the cell lines, 7-day L1210 ascites, maintained by transfer in mice, were the most sensitive. The greater sensitivity of these cells, as compared to 4-day L1210 ascites or cells in culture, may be explained by the difference in their growth stages. The 7-day ascites cells would be in stationary growth as compared to the exponentially growing 4-day ascites or the cells in culture. The temperature sensitivity of Chinese hamster ovary cells in different parts of the cell cycle was determined. Mid and late-S-phase cells were more sensitive than cells in mitosis or early S; G1 and G2 cells were the least sensitive. In a partially synchronized culture, the heat sensitivity of L1210 cells increased with increased percentage of cells in S phase.