USE OF 2,3-NAPHTHALENEDICARBOXALDEHYDE DERIVATIZATION FOR SINGLE-CELLANALYSIS OF GLUTATHIONE BY CAPILLARY ELECTROPHORESIS AND HISTOCHEMICAL-LOCALIZATION ION BY FLUORESCENCE MICROSCOPY

We report that 2,3-naphthalenedicarboxaldehyde reacts rapidly with glu tathione and its precursor, gamma-glutamylcysteine, to form highly flu orescent derivatives under physiological conditions. In contrast to pr evious accounts of 2,3-naphthalenedicarboxaldehyde labeling of primary amines, no additional CN- ion or any other additional nucleophile is required. The fluorescence spectral properties of the chromophores (ga mma(exc max) = 472 nm, lambda(em max) = 528 nm) make these derivatives amenable to excitation and detection by optical instrumentation that is optimized for fluorescein wavelengths. This selective labeling chem istry enabled quantitative determination and histochemical localizatio n of glutathione in neurobiological samples, Intracellular glutathione was labeled by incubating cultured cells or cell suspensions in a 2,3 -naphthalenedicarboxaldehyde-supplemented, DMSO-containing physiologic al buffer (pH = 7.4) for 2-10 min. Applications include imaging of cul tured NG 108-15 cells (mouse neuroblastoma x rat glioma) and primary g lial and neuronal cell cocultures (rat hippocampus) using epiluminesce nt and confocal fluorescence microscopy. Quantita tive determination o f glutathione in single NG 108-15 cells was accomplished using laser-i nduced fluorescence detection and capillary electrophoresis.