PMID: 8450203Apr 1, 1993Paper

A photocross-linking fluorescent indicator of mitochondrial membrane potential

The Journal of Histochemistry and Cytochemistry : Official Journal of the Histochemistry Society
K M HahnA S Waggoner

Abstract

Ionic dyes that distribute across membranes according to electrical potential have proven valuable as fluorescent indicators of mitochondrial energetics in living cells. Applications have been limited, however, as potential-dependent staining is lost during cell fixation. We have produced a membrane potential indicator whose potential-dependent distribution can be made permanent, to enable correlation of membrane potential with cytochemical information from immunofluorescence. A carbocyanine dye was derivatized with a photoreactive nitrophenylazide moiety so that irradiation would induce nonspecific, covalent attachment to nearby molecules. Photo-induced cross-linking was observed in paper chromatography, when irradiation caused immobilization of the dye. The new dye, named PhoCy (photofixable cyanine), showed specific staining of mitochondria in living Swiss 3T3 fibroblasts. When living cells were stained, irradiated, and fixed with formaldehyde, mitochondrial staining was retained owing to cross-linking with mitochondrial components. Omission of irradiation eliminated mitochondrial staining in fixed cells. Labeling, irradiation, and fixation procedures were optimized to produce bright specific staining with minimal background...Continue Reading

Citations

Dec 1, 1993·Journal of Fluorescence·B Ehrenberg, L M Loew
Apr 1, 1996·Journal of Photochemistry and Photobiology. B, Biology·J Plásek, K Sigler
Jun 30, 2000·Methods : a Companion to Methods in Enzymology·H M Shapiro

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