In toddlers, the transition from home to childcare might elicit high levels of the stress hormone cortisol. Measuring cortisol may give an indicator for children's experience and hence, may help improve this transition. We applied linear mixed model analyses to investigate the cortisol levels of 119 toddlers during their transition to childcare across time of day (morning, afternoon, and evening) and phase (accompanied by parents, separated from parents, and after four to six weeks in childcare). The influence of age, gender, number of siblings, and childcare group size was analyzed. Time of day and phase influenced cortisol levels significantly. On average, children had elevated cortisol levels in the afternoon throughout transition, with the peak coming in the separation phase. Cortisol levels declined significantly toward the evening. Children younger than 14 months showed higher evening levels and higher afternoon levels after 4-6 weeks in childcare. The findings suggest that the onset of childcare -- particularly separation from parents -- may be demanding for toddlers. Low evening levels indicate relief of tension at home. Higher levels of afternoon cortisol of under 14-months-old children at the follow-up measurement may indicate that younger children need more time to settle in at childcare.