Electrochemical sensors for the determination of cholesterol with Au as a working electrode (Au) and its application to the analysis of urine have been done. Gold electrode was preparation using gold pure (99.99%), with size 1.0 mm by length and wide respectively, connected with silver wire using silver conductive paint. Validation methods have been investigated in the analysis of cholesterol in human urine using electrochemical sensors or cyclic voltammetry (CV) method. The effect of electrolyte and uric acid concentration has been determined to produce the optimum method. Validation method parameters for cholesterol analysis in human urine using CV are precision, recovery, linearity, limit of detection (LOD) and limit of quantization (LOQ). The result showed the effect of concentration on current with R²= 0.991. The results of the validation method showed the precision, recovery, linearity, LOD and LOQ are 1.2539%, 144.33%, 0.991, 1.49 x 10^-4 M and 4.96 x 10^-4 M, respectively. As a conclusion is Au electrode is a good electrode for electrochemical sensors to determination of cholesterol in human urine.