Background: The most frequent cause of respiratory failure in preterm newborns, as well as the most frequent cause of mortality and long-term morbidity associated with prematurity, is respiratory distress syndrome (RDS). RDS affects half of newborns with birth weights (BW) under 1.5 kg, and a sizable portion of those newborns also experience acute kidney injury (AKI). Objective: The purpose of this study was to identify useful biomarkers (serum cystatin C andNeutrophil gelatinase-associated lipocalin (NGAL) to predict AKI in premature infant with RDS. Patients and Methods: This was a prospective (case-control) study was conducted on 90 preterm neonates between 28 and 36 gestational weeks (GW) from the neonatal intensive care unit (NICU) of Benha University hospitals.  Serum creatinine, Blood urea nitrogen Levels, Serum cystatin C and Serum NGAL were measured for all included neonates.  Results: Serum creatinine showed no significant difference between the studied groups at day 3 (P = 0.273). At day 5 and day 7, it was significantly higher in group I (1.4 & 1.8 mg/dl, respectively) than groups II (0.7 & 0.6 mg/dl, respectively) and III (0.6 & 0.5 mg/dl, respectively). Also, it was significantly higher in group II (0.7 & 0.6 mg/dl, respectively) than group III (0.6 & 0.5 mg/dl, respectively). Serum cystatin C and serumNGAL showed an overall significant difference between the studied groups at day 3 (P < 0.001). In post hoc analyses, it was significantly higher in group I (1.7 mg/l) than groups II (1.1 mg/l) and III (01 mg/l). Conclusion: NGAL and sCys C levels were found to have a statistically significant association with development of AKI in preterm neonates with RDS and they were elevated earlier than sCr which makes NGAL and sCys C a good predictive marker for AKI in preterm neonates better than sCr.