Flow cytometric analysis of intracellular pH in cultured opossum kidney (OK) cells |
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Authors: | Marshall H. Montrose Marc A. Condrau Heini Murer |
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Affiliation: | (1) Physiology Institute, University of Zurich, 8057 Zurich, Switzerland;(2) Institute for Biomedical Engineering, University of Zurich and ETH, 8044 Zurich, Switzerland;(3) Present address: Johns Hopkins University, Huntarian 515, 725 N. Wolfe St., 21205 Baltimore, MD |
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Abstract: | Summary Suspensions of OK cells (a continuous renal epithelial cell line originating from the opossum kidney) were examined by flow cytometry. Three parameters were evaluated simultaneously; cell integrity as assayed by propidium iodide fluorescence, cell size as measured by time-of-flight, and intracellular pH as measured by fluorescence of 2,7-bis-(2-carboxyethyl)-5,6 carboxyfluorescein (BCECF). The suspension was shown to be composed of both intact singlets and doublets of cells, and no difference was noted in the behavior of these two populations with respect to the resting intracellular pH, or of the response of intracellular BCECF to changes in pH. Evidence suggests that using NH4 prepulses to create an acid load broadens the intracellular pH distribution. The population of OK cells demonstrates a recovery from this acid load which is very homogeneous with respect to its sensitivity to Na+ removal or EIPA (ethylisopropyl-amiloride), suggesting that virtually all cells utilize Na+/H+ exchange for this recovery. The data also suggest heterogeneity in the cellular pH recovery from an acid load with respect to the observed rates of Na+/H+ exchange. Despite this heterogeneity, the Na+/H+ exchanger is observed to focus the resting intracellular pH of the population to approximately pH 7.4–7.5. The response of the population to PTH suggests that the majority of cells respond to the hormone, and that the total Na+/H+ exchange in individual cells is only partially inhibited even in the presence of saturating PTH concentrations. |
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Keywords: | intracellular pH BCECF propidium iodide time-of-flight epithelial cell culture opossum kidney |
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