An aqueous solution of Nile Red (NR)-absorbed 32-nm gold nanoparticles (AuNPs) have been used to sense glutathione (GSH). When the NR product is displaced by GSH on the AuNP surface, the fluorescence of the solution increases and the AuNPs aggregate. To determine the concentration and distribution of GSH within erythrocyte cells, a homemade fluorescence and scattering microscope was constructed. This system allows monitoring, within individual cells, of the uptake and transportation of the NRAuNPs and the displacement of the NR product from the NRAuNP surface by GSH. The fluorescence and scattering images clearly indicate the location of GSH inside the cells; these findings are supported by images recorded using 2,3-naphthalenedicarboxaldehyde, which is a highly selective fluorogenic reagent for GSH. Microscopic fluorescence measurements of the NRAuNPs revealed that the GSH concentration inside erythrocyte cells is 1.30 +/- 0.31 mM. To confirm this result, lysed erythrocyte cells were analyzed by applying capillary electrophoresis in conjunction with laser-induced fluorescence using NRAuNPs; accordingly, the average GSH concentration in a single erythrocyte cell was determined to be 1.32 +/- 0.06 mM.