A fluorescence-microscopic and flow-cytometric study of HeLa cells with an experimentally induced respiratory deficiency.
HeLa cells, cultured over a long period in a medium containing low doses of ethidiumbromide, were used as a model system for flow-cytometric detection of human cells with impaired mitochondrial respiratory function. Based on laserscan and flowcytometric analysis after rhodamine 123 staining, the mitochondrial membrane potential of respiratory deficient cells seems unchanged as compared to control cells. Maintenance of this membrane potential in respiration-impaired cells requires glycolytic ATP generation, as transient inhibition of glycolysis by sodium fluoride affects rhodamine 123 accumulation in ethidiumbromide-treated cells, but not in control cells. We present a protocol which allows the detection and separation of respiratory deficient cells by flow cytometry.
Skowronek
P
P
Institut für Pathologie, Universität Ulm, Germany.
Haferkamp
O
O
Rödel
G
G
D000255:Adenosine Triphosphate / Q000378:metabolism; D002469:Cell Separation; D004272:DNA, Mitochondrial / Q000378:metabolism; D004734:Energy Metabolism; D004996:Ethidium / Q000494:pharmacology; D005260:Female; D005434:Flow Cytometry; D006019:Glycolysis; D006367:HeLa Cells; D006801:Humans; D008564:Membrane Potentials; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism*; D010101:Oxygen Consumption / Q000187:drug effects; D020112:Rhodamine 123; D012235:Rhodamines / Q000378:metabolism; D012969:Sodium Fluoride / Q000494:pharmacology
D004272:DNA, Mitochondrial; D012235:Rhodamines; D020112:Rhodamine 123; D000255:Adenosine Triphosphate; D012969:Sodium Fluoride; D004996:Ethidium