Patients developing acute kidney injury (AKI) after cardiac surgery have increased mortality compared with those with stable postoperative renal function. A reliable marker of early AKI in serum or urine has not been found. To test the hypothesis that urinary cystatin C (CyC) increases with greater severity of AKI, we prospectively studied 45 patients undergoing cardiac surgery. Perioperatively, blood and urine were sampled for standard chemistries and CyC (measured by ELISA). AKI was defined as > 25% increase in serum creatinine or the initiation of renal replacement therapy (RRT) within the first 3 postoperative days. AKI developed in 24 patients (53.33%), 5 of whom received RRT (11.1% of the total). The 3-day postoperative maximum increase of urinary CyC from the preoperative baseline was significantly different via ANOVA in those receiving RRT (n = 5) from those with AKI without RRT (n = 16) and those with no AKI (n = 12) (p = .033). This effect is present for the first four postoperative samples, which encompass the first 48 postoperative hours. There was no statistical difference in the preoperative (baseline) urine CyC values. Furthermore, the maximum 3-day increase in perioperative urinary fractional excretion of CyC (FECC) was significantly different across all three groups (no AKI, AKI without RRT, and RRT), p = .01. Similar to urinary CyC, this difference was seen for the first 48 postoperative hours. There was no statistical significance to the preoperative values across all three groups. Increased urinary cystatin C excretion in the early postoperative period precedes the development of severe AKI (requiring RRT) and correlates with AKI severity. Urinary CyC and FECC are promising biomarkers of AKI following adult cardiac surgery.