Background:The neonatal period is the most vulnerable time of human life for diseases. Neonatal morbidity and mortality are significant contributors to under-five morbidity and mortality in sub-Saharan Africa. Objective: To assess prevalence and factors associated with neonatal mortality at Ayder Comprehensive Specialized Hospital. Methods: A facility-based cross-sectional study was conducted on neonates admitted to the neonatal intensive care unit of Ayder Comprehensive Specialized Hospital from June 1, 2018 to May 30, 2019. Data were taken retrospectively from patient records during admission, discharge, and death certificate issue. The data were entered and analyzed using SPSS version 23. Descriptive and logistic regression analysis was done to describe and identify associated factors with neonatal mortality. P-values <0.05 were considered statistically significant. Results: During the study period, 1785 neonates were seen and 1069 (60%) were males. Neonatal mortality rate was 298 (16.7%). Of all the deaths, 98.3% occurred during the first 7 days of age. Respiratory distress syndrome (AOR: 12.56; 95% CI: 6.40-24.66:), perinatal asphyxia (AOR: 19.64; 95% CI: 12.35-31.24), congenital anomaly (AOR: 2.42; 95% CI: (1.48-4.01), early neonatal sepsis (AOR: 3.68; 95% CI: 2.32-5.81), late onset sepsis (AOR: 8.9; 95% CI: 4.14-19.21), gestational age, 34-36+6 weeks (AOR: 0.09; 95% CI: 0.014-0.59), 3741+6 weeks (AOR: 0.025; 95% CI: 0.0030.218), >42 weeks (AOR: 0.039; 95% CI: 0.004-0.4250), parity (AOR: 0.64; 95% CI: 0.44-0.93) and hospital stay (AOR: 0.09; 95% CI: 0.05-0.14) were significantly associated with neonatal mortality. Neonates with a birth weight of less than 1500 g were at 49%, 70%, and 80% increased odds of mortality compared to those 1500-2499 g, 2500-3999 g, and more than 4000 g, respectively. Conclusion: In this study neonatal mortality was significantly high. Neonatal mortality was highly associated with primipara, prematurity, low birth weight, perinatal asphyxia, respiratory distress syndrome, congenital anomaly, neonatal sepsis and duration of hospital stay. Many of those cases could be prevented by improving antenatal care follow up, emergency obstetric services, and the enhancement of neonatal resuscitation skills and management of sick neonates.