The use of personal protective equipment (PPE) in prehospital emergency care has significantly increased since the onset of the coronavirus disease 2019 (COVID-19) pandemic. Several studies investigating the potential effects of PPE use by Emergency Medical Service providers on the quality of chest compressions during resuscitation have been inconclusive. This study aimed to determine whether the use of PPE affects the quality of chest compressions or influences select physiological biomarkers that are associated with stress. This was a prospective randomized, quasi-experimental crossover study with 35 Emergency Medical Service providers who performed 20 minutes of chest compressions on a manikin. Two iterations were completed in a randomized order: (1) without PPE and (2) with PPE consisting of Tyvek, goggles, KN95 mask, and nitrile gloves. The rate and depth of chest compressions were measured. Salivary cortisol, lactate, end-tidal carbon dioxide (EtCO2), and body temperature were measured before and after each set of chest compressions. There were no differences in the quality of chest compressions (rate and depth) between the two groups (P >.05). After performing chest compressions, the group with PPE did not have elevated levels of cortisol, lactate, or EtCO2 when compared to the group without PPE, but did have a higher body temperature (P <.001). The use of PPE during resuscitation did not lower the quality of chest compressions, nor did it lead to higher stress-associated biomarker levels, with the exception of body temperature.