Selective detection of endogenous thiols using microchip-based flow analysis and mercury/gold amalgam microelectrodes

This paper describes the fabrication and characterization of thin-layer mercury/gold amalgam microelectrodes and their integration with microchip-based flow injection analysis. This microchip platform allows on-chip injection and lysis of erythrocytes followed by selective detection of intracellular glutathione (GSH) at low potentials. The thin-layer gold microelectrodes were amalgamated by electrodeposition of mercury. The electrodes produced a linear response for both GSH and cysteine in flow injection analysis studies utilizing both off-chip and on-chip injection. Comparative experiments using diamide and on- chip injection were performed to demonstrate the ability of the microchip device to detect changes in GSH concentration. Finally, rabbit erythrocyte samples (2% hematocrit) were injected and lysed on- chip and the amount of GSH detected corresponded to 312 amol/cell, which is in agreement with previously reported values. The selectivity, short time between injection and detection (similar to 5 s), and the continuous introduction of sample to the on- chip injector should enable the study of dynamically changing systems such as the glutathione redox system found in erythrocytes.