Abstract Background: Patient safety has become a priority in healthcare system and among health care workers. Nurses play a major role in patients' safety through twelve hour shift because they are accountable for direct and continuous patient care. So errors occur for a variety of reasons, including the effects of twelve hours shift. It is estimated that 1.3 million health care errors occur each year and of those errors 48,000 to 98,000 result in the deaths of patients. Aim of Study: To assess the relationship between twelve hour shift and patient safety. Subjects and Methods: A descriptive research design was utilized to collect data from (150) staff nurses working in all Medical, Surgical Departments and Intensive Care Units at National Cancer Institute which is affiliated to Cairo University having at least one year of experience. Data was collected by using Hospital Survey of Patient Safety Culture (HSOPSC) and patient safety assessment manual. Results: The study revealed that the dimension with the highest positive responses was; teamwork within units (mean= 4.61) and the lowest positive response was in the dimension of non-punitive response to error (mean=1.81) and also feedback and communication about error (mean=2.71). Conclusion: The study concluded that there was a signif-icant relation between 12 hour shifts and patient safety dimen-sions. Recommendations: The study recommended Schedule sensibly for nurses to decrease fatigue that leading to error. Head nurses should be encouraged to establish non-punitive environment as well as a teamwork spirit among nursing staff working in critical care and general ward staff nurses. Develop a well-established system for incident reporting, and all staff nurses should be informed and trained about it.