Literature evaluating intraoperative temperature/humidity and risk of surgical site infection (SSI) is lacking. All operations at three centers reported to the ACS-NSQIP were reviewed (2016–2020); ambient intraoperative temperature (⁰F) and relative humidity (RH) were recorded in 15-min intervals. The primary endpoint was superficial SSI, which was evaluated with multi-level logistic regression. 14,519 operations were analyzed with 179 SSIs (1.2%). The lower/upper 10th percentiles for temperature and RH were 64.4/71.4 °F and 33.5/55.5% respectively. Low or high temperature carried no significant increased risk for SSI (Low ⁰F OR = 0.95, 95% CI 0.51–1.77, P = 0.86; High ⁰F OR = 1.13, 95% CI = 0.69–1.86, P = 0.63). This was also true for low and high RH (Low RH OR = 0.96, 95% CI 0.58–1.61, p = 0.88; High RH OR = 0.61, 95% CI = 0.33–1.14, P = 0.12). Analysis of combined temperature/humidity showed no increased risk for SSI. Significant deviations in intraoperative temperature/humidity are not associated with increased risk of SSI. •Large deviations in OR temperature do not increase risk of SSI.•Large deviations in OR humidity do not increase risk of SSI.•Efforts to decrease risk of SSI should focus on other modifiable factors.