To examine the influence on postoperative endothelial cell density (ECD) caused by preoperative temperature change from 4 °C to room temperature in donor corneas for Descemet stripping automated endothelial keratoplasty (DSAEK). This retrospective, case-controlled comparison of 100 eyes that underwent DSAEK using imported donor corneas transferred from an overseas eye bank (SightLife, Seattle, WA, USA). Fifty donor corneas experienced temperature reversal for ECD measurement (TR group), and postoperative outcomes were compared with 50 disease-matched cases that did not experience temperature changes before DSAEK (NTR group). The main outcome measure was endothelial cell loss and reduction rate of ECD at 1, 3, and 6 months following DSAEK. ECD at 3 months following DSAEK was significantly less in the TR group (1458 ± 494/mm2) than in the NTR group (1696 ± 374/mm2; P = 0.014), though ECD at 6 months was not significantly less in the TR group. The reduction rate of ECD at 3 months was greater in the TR group (42.3% ± 17.2%) than in the NTR group (35.7% ± 14.2%; P = 0.044), though reduction rate of ECD at 6 months was not significantly less in the TR group. We found that preoperative temperature change in donor corneas may adversely affect ECD following DSAEK.