Hydroquinone diphosphate/Ag+ as an enzymatic substrate for alkaline phosphatase catalyzed silver deposition

Signal amplification by enzyme labels such as alkaline phosphatase (AP) used in affinity assays (immuno or DNA based assays) is a usual strategy to get high sensitive detection methodologies. An advantageous combination of an enzyme substrate Hydroquinone diphosphate (HQDP) with silver ions (Ag+) in solution is presented in this work as a new enzymatic substrate for AP. This enzyme catalyzes the dephosphorylation of HQDP producing hydroquinone, a reducing agent for silver ions present in solution. Thus by mixing AP, HQDP and Ag+, metallic silver is deposited where the enzymatic reaction takes place. A qualitative assay is presented in nitrocellulose membranes to demonstrate this mechanism. The affinity reaction between streptavidin and biotin (labeled with AP) is also followed by the electrochemical detection of silver deposited on the surface of Screen Printed Carbon Electrodes. Anodic stripping voltammetry of enzymatically deposited silver is showed as a sensitive detection technique for biosensing applications.