This study aimed to assess subjective and objective parameters of stress among nurses during the COVID-19 pandemic and to examine the recovery effect of a day off. In this prospective observational trial, we measured heart rate variability (using a wearable device) and perceived stress levels on 3 working days and 1 day off. We obtained the following data using an online questionnaire: working conditions, COVID-19-related problems, depression (Patient Health Questionnaire-9), anxiety (Generalized Anxiety Disorder-7), effort-reward imbalance, and work-family conflict in a sample of German nurses (N = 41). When comparing working days with a day off, we observed a significant difference for physical load (Cohen's d = 0.798, P < .001), mental load (Cohen's d = 0.660, P = .001), emotional exhaustion (Cohen's d = 0.945, P < .001), and overburdening (Cohen's d = 0.585, P = .002) with higher scores on working days. Regarding heart rate variability, we did not find a difference. Correlational analyses revealed a significant association between being afraid to get infected with COVID-19 and lower heart rate variability (r = -0.336, P = .045) and between being afraid to infect relatives and lower heart rate variability (r = -0.442, P = .007). Furthermore, a higher total sum score of work-family conflict was significantly associated with lower heart rate variability (r = -0.424, P = .01). As heart rate variability observations were different from those regarding subjectively perceived stress, further studies are needed to evaluate and differentiate the influence of work stress and other types of stress on heart rate variability.