Introduction: With the advancement of various industries, different types of shift work are being implemented throughout the world. Shift work is considered necessary to ensure continuity of care in hospitals and residential facilities. In particular, the night shift is one of the most frequent reasons for the disruption of circadian rhythms, causing significant alterations of sleep and biological functions that can affect physical and psychological well-being and negatively impact work performance. The aim of this study was to compare between fatigue, sleep disturbance and job satisfaction among nursing staff working in shift duties and day work in tertiary care hospital in Lahore, Pakistan. Methods: A quantitative analytical cross-sectional study was conducted to compare fatigue, sleep disturbance and job satisfaction among shift duties and day work among nurses in tertiary care hospital in Lahore, Pakistan. The descriptive statistics include the demographic data, factors and represented using the frequencies and percentage. To find out the relationship between the factors Pearson correlation was applied. Population of the study was nurses and sample size is 350 participants. Results: A significant positive correlation was found between the level of fatigue and the levels of anxiety and depression. In our study; the findings indicated that the level of job stress is moderate among day rotation nurses whereas it was severe. There is a positive relationship between marital life and fatigue and sleep disorders. Conclusion: In this study it was identified that nurses working in the shift duties have poor sleep quality. Whereas the nurses working in the day shifts have good sleep quality. Participants also reported that harsh interactions with other interdisciplinary team members worsen the fatigue and job satisfaction among the nurses. Key words: Shift Duties, Job Satisfaction, Fatigue, Sleep Disorder.