A miniaturised electrochemical affinity assay based on a wall-free sample droplet and micro-dispensing of the redox-labelled binding partner

An affinity-assay was developed that is based on the modulation of the diff usion coefficient of a redox-labelled hapten upon complementary recognition of the analyte leading to an increase of molecular weight and hence to a d ecrease of the diffusion coefficient. The slower diffusion is monitored by means of cyclic voltammetry. In order to demonstrate the feasibility of thi s assay format, recognition of biotin by streptavidin has been chosen as a model system. Labelling of biotin was achieved by covalent binding of a fer rocene derivative to the biotin unit. To reduce the consumption of expensiv e compounds and to allow automatisation of the assay a novel miniaturised s et-up was developed based on a wall-free sample droplet which forms the ele ctrochemical cell with typical volumes of up to 10 mul. This droplet is dis pensed by means of a step-motor driven syringe pump through a specially des igned electrode holder spanning the gap between a micro-working electrode a nd a macroscopic counter electrode. By means of a piezo-driven micro-dispen ser a predefined number of nano-droplets (100 pl volume each) containing th e redox-labelled hapten are shot into the sample droplet. By this, any phys ical contact and hence any cross-contamination between the sample and the r eagent solution could be avoided. Signal amplification can be achieved by r edox recycling between the micro-electrode and the perpendicular positioned macroscopic counter electrode. (C) 2001 Elsevier Science B.V. All rights r eserved.