The enzymatic method for the determination of serum creatinine is accepted as one of the most accurate methods in a clinical laboratory. This method was used on a biochemistry auto analyzer (Cobas Integra 400) to determine serum creatinine at the laboratory of University Hospital – Pleven. The characteristics and reliability of this enzymatic method for creatinine were compared with the Jaffe kinetic method. Effects of some interfering substances like bilirubin and glucose on the Jaffe kinetic method and the enzymatic method were compared. Glucose and bilirubin inhibit the reaction between creatinine and alkaline picrate. Glucose slowly reduces picric acid to picrate, while the bilirubin present in a sample is oxidized to biliverdin under alkaline conditions. This leads to a decrease in absorbance at 520 nm. We measured creatinine in serum samples with the enzymatic method and the Jaffe kinetic method in samples divided into four groups: group I –samples without bilirubin and glucose ; group II –samples with high level of glucose; group III - samples with high level of bilirubin, group IV – all samples. For Group I, the correlation coefficient obtained by comparing the enzymatic creatinine method and Jaffe's kinetic method was R = 0.983. There was a very good agreement between the two methods in terms of correlation coefficient even in the samples with high levels of glucose or bilirubin.