Catalysis-electrochemical determination of zeptomole enzyme and its application for single-cell analysis

A novel electrochemical method for determination of zeptomole amounts of enzyme was developed by a combination of on-capillary enzyme-catalyzed reaction and electrochemical detection. A limit of detection (LOD) of zeptomole (zmol, 10(-)(21) mol) was achieved by monitoring the product of the enzyme-catalyzed reaction. In this method, after enzyme molecules were electrokinetically injected into the capillary, they were electromigrated to the section of the capillary immersed in a warm water bath, where the enzyme molecules reacted with the enzyme substrates in the running buffer in the presence of the activator of the enzyme-catalyzed reaction. Then the electroactive product zone of the enzyme-catalyzed reaction was electromigrated to the horn-shaped outlet of the capillary and electrochemically detected by a carbon fiber disk bundle electrode at a constant potential. Glucose-6-phosphate dehydrogenase (G6PDH) was chosen as the model enzyme. A LOD of 1.3 zmol was achieved. This method was applied to determine zeptomoles of G6PDH in individual human erythrocytes.