An electrochemical amplification immunoassay using biocatalytic metal deposition coupled with anodic stripping voltammetric detection

An electrochemical amplification immunoassay is reported using biocatalytic metal deposition coupled with anodic stripping voltammetric detection. In this method, the captured antibody was first immobilized onto a gold electrode via a self-assembled layer. After a sandwich immunoreaction, alkaline phosphatase-labeled antibody was bound to the gold electrode. The alkaline phosphatase on the electrode catalyzes the hydrolysis of ascorbic acid 2-phosphate to produce ascorbic acid. The latter, in turn, reduced silver ions on the electrode surface, leading to the deposition of silver onto the protein-modified electrode surface. The deposited metal was electrochemically stripped into solution and then measured by anodic stripping voltammmetry. Compared with the direct voltammetric detection of ascorbic acid, anodic stripping voltammetric detection of metal ions is more sensitive. For the amount of deposited silver relates to the amount of enzyme-generated ascorbic acid, which was controlled by the amount of enzyme bound on the electrode surface, the stripping current signal reflects the amount of target protein, achieving a linearly relationship in the range from 5 to 1000 ng mL−1 in a logarithmic plot with a detection limit of 2.2 ng mL−1. The utilization of the high biocatalytic activity of enzyme and the sensitive anodic stripping voltammetry to detect metal ions dramatically enhanced the sensitivity in immunoassay.