The objective of this study was to evaluate the effects of heat stress and nighttime temperature drop on oocyte yield and in vitro embryo production (IVEP) success. The study was conducted on Holstein heifers within the same herd, under similar management and feeding conditions. Each session of ovum pick-up (OPU) was classified into continuous heat stress (CHS; THImax ≥72 and THImean ≥68), high daytime temperature–humidity index (THI) with nighttime recovery (HDT-NR; THImax ≥72 and THImean <68), and thermoneutral (TN; THImax <72 and THImean <68) groups based on environmental THI values. The TN group was found to have a greater number of small- to medium-sized follicles and a higher total number of follicles in comparison to the CHS group (p<0.05). In terms of oocyte yield, the numbers of grade-A, total, and viable oocytes were found to be higher in the TN and HDT-NR groups than in the CHS group (p<0.05). The assessment of the IVEP revealed that the mean number of blastocysts per OPU was lower in the CHS group (p<0.05). Consequently, even when the daytime maximum THI was high, nighttime cooling, reflected by a lower mean THI, appeared to preserve OPU and IVEP performance without adverse effects. Therefore, during the summer season, when the likelihood of heat stress is increased, considering nighttime recovery while planning and conducting in vitro embryo production processes may be an important criterion for improving production success.