Abstract: A new analytical system by using bromothymol blue (BTB) as an electrochemical probe in B-R buffer solution of pH 3.5 for determination of protein was proposed.A sensitive voltammetric reductive peak of BTB was observed at -0.458 V (vs.SCE).Upon the addition of human serum albumin (HSA) to the reaction system,the peak potential of BTB was not changed but the peak current was decreased.It was found that linear relationship between the 2nd derivative of the magnitude of decrease in peak current (ΔI″p) and the increase of mass concentration of HSA in the range within 40.0 mg·L-1 as shown by the linear regression equation: ΔI″p=133.52 C-3.72 (r=0.999) was obtained.Determination limit of this reaction for HSA was found to be 1.0 mg·L-1.Combination ratio of HSA to BTB was found to be 1∶4,and the combination constant to be 1.43×1019.The proposed method was used in the determination of HSA in blood samples of healthy persons,giving results in consistency with the results obtained by the conventional Comassie brilliant blue G-250 photometric method.This method was also applied to determined other kinds of protein,such as bovine serum albumin (BSA),bovine hemoglobin (BHb),ovalbumin (OVA) and etc.