Prehospital advanced life support (ALS) has been offered in many countries for patients experiencing out-of-hospital cardiac arrest (OHCA); however, its effectiveness remains unclear. This study aimed to determine the impact of emergency medical service (EMS) with ALS training as a nationwide pilot project for adults with OHCA in the Republic of Korea. This retrospective multicenter observational study was conducted between July 2019 and December 2020 using the Korean Cardiac Arrest Research Consortium registry. The patients were categorized into an intervention group that received EMS with ALS training and a control group that did not receive EMS with ALS training. Conditional logistic regression analysis was performed using matched data to compare clinical outcomes between the two groups. Compared with the control group, the intervention group had a lower rate of supraglottic airway usage (60.5% vs. 75.6%) and a higher rate of undergoing endotracheal intubation (21.7% vs. 6.1%,P< 0.001). In addition, the intervention group was administered more intravenous epinephrine (59.8% vs. 14.2%,P< 0.001) and used mechanical chest compression devices more frequently in prehospital settings than the control group (59.0% vs. 23.8%,P< 0.001). Based on the results of multivariable conditional logistic regression analysis, survival to hospital discharge (odds ratio: 0.48, 95% confidence interval: 0.27–0.87) of the intervention group was significantly lower than that of the control group; however, good neurological outcome was not significantly different between the two groups. In this study, survival to hospital discharge was worse in patients with OHCA who received EMS with ALS training than in those who did not.