Monitoring an oxidative stress mechanism at a single human fibroblast

Easily oxidizable substances inside human diploid fibroblast cell strains were monitored amperometrically with a platinized carbon-fiber microelectrode. The experiment involved positioning a microelectrode over a single biological cell, forcing the electrode tip into the cell via micromanipulator control, and measuring the transient current corresponding to the complete electrolysis of electroactive species released by the cell. A second series of experiments involved puncturing a hole into the cell with a micropipet and measuring the transient current corresponding to the complete electrolysis of electroactive species emitted by the cell with an electrode positioned above the cell. The selectivity of both amperometric measurements was demonstrated through the use of known hydrogen peroxide scavengers (added catalase or intracellular peroxidase + added o-dianisidine) to the media bathing the cells. The abolition of the amperometric signal under these conditions suggested that hydrogen peroxide was the primary substance detected. The magnitude and the time course of the transient current measured implied that the hydrogen peroxide detected was not only that initially present in the cell before its membrane was pierced but represented mostly an oxidative stress response of the cell to its injury.