Neonatal infections are a leading cause of morbidity and mortality, necessitating widespread antibiotic use in Neonatal Intensive Care Units (NICUs). This study examines antibiotic prescription patterns in the NICU at District Civil Hospital Nuh Mandikhera, , between April 2023 and April 2024, through a prospective observational analysis of 50 neonates. The study evaluates antibiotic utilization, class distribution, duration, combination therapies, and adherence to national prescribing guidelines. Findings reveal a high antibiotic prescription rate of 100%, with an average of 3.02 antibiotics per neonate, predominantly involving aminoglycosides (amikacin), cephalosporins (cefotaxime), and carbapenems (meropenem). Combination therapies, particularly piperacillin with amikacin, were frequently prescribed, with limited reliance on bacterial culture data, as only 2% of cases yielded positive results. While 70.5% of prescriptions complied with the National Essential Drug List (NEDL), generic prescribing was entirely absent (0%), and prolonged therapy durations with a mean of 14.6 days further raised concerns about antimicrobial resistance. These findings underscore the urgent need for an NICU-specific antibiotic stewardship program to optimize empirical therapy, encourage microbiology-guided prescribing, enhance adherence to national guidelines, and mitigate the risk of antimicrobial resistance. Future multi-center studies are recommended to further validate these findings and guide policy interventions in neonatal antimicrobial management.