We describe a procedure for specific, rapid, kinetic determination of creatinine, in which a manual coupled-enzyme micro-scale assay is adapted to a centrifugal analyzer. The creatinine reaction is ultimately linked to NADH utilization, which is measured by the absorbance change at 340 nm. This procedure requires 15 microL of serum and the standard curve is linear to a creatinine concentration of 200 mg/L. A four-point kinetic algorithm allows the dynamic range of the assay to be extended without sacrificing sensitivity, and makes a separate serum blank unnecessary. The within-run precision (CV) for samples with a creatinine concentration of 11 and 52 mg/L was 5.6 and 2.4%, respectively; day-to-day CV for a creatinine concentration of 11 mg/L was 7.7% (n = 21). We compared this procedure with a kinetic Jaffé procedure, with excellent agreement (r = 0.996; y = 0.96x + 2.4 mg/L). Bilirubin, non-esterified fatty acids, and ketone bodies do not affect creatinine determinations by this method; thus the method is especially useful for monitoring the renal function of diabetics.