Direct evaluation of fluorescence in single renal epithelial cells using a mitochondrial probe (DASPMI)

The study of distribution and quantitation of a fluorescent probe in living epithelia with the aid of an inverted microscope requires that individual cells can be analysed without optical interference from adjacent cells. This report describes the application of fluorescence microscopy and fluorometry to a recently developed in vitro culture system of renal epithelial cells. Epithelial cells derived from the mammalian renal cortical collecting tubule (CT) and the thick ascending loop of Henle (TAL) are cultivated as continuous monolayers in serum-free, hormone-supplemented media. A specific mitochondrial marker (DASPMI) is added to the medium and incorporated into the cytoplasm. The microscopic image reveals that the mitochondrial fluorescence distribution differs between CT and TAL cultures. The fluorometric quantitation shows a normally distributed histogram of medium-range intensity in TAL cell cultures while CT cultures exhibit a two-peak pattern of mitochondrial fluorescence distribution among epithelial cells.