Objectives Night shift work has been associated with poor sleep, weight gain, metabolic syndrome, which are recognised risk factor for diabetes. However, only a few studies have examined the effect of shift work on diabetes risk. Here, we study the association between shift work and incidence of diabetes in Danish nurses. Methods We used the Danish Nurse Cohort with 28 731 participating female nurses recruited in 1993 (19 898) or 1999 (8833), when self-reported baseline information on diabetes prevalence, lifestyle and working time were collected, and followed them in the Danish Diabetes Register for incidence of diabetes until 2013. Nurses reported whether they worked night, evening, rotating or day shifts. We analysed the association between working time and diabetes incidence using a Cox proportional hazards model adjusted for diabetes risk factors, separately with and without adjustment for body mass index (BMI) which might be an intermediate variable. Results Of 19 873 nurses who worked and were diabetes-free at recruitment, 837 (4.4%) developed diabetes during 15 years of follow-up. The majority of nurses (62.4%) worked day shifts, 21.8% rotating shift, 10.1% evening and 5.5% night shifts. Compared with nurses who worked day shifts, we found statistically significantly increased risk of diabetes in nurses who worked night (1.58; 1.25 to 1.99) or evening shifts (1.29; 1.04 to 1.59) in the fully adjusted models including BMI. Conclusions Danish nurses working night and evening shifts have increased risk for diabetes, with the highest risk associated with current night shift work.