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Metachromatic Differential Fluorochroming of Living and Dead Ascites Tumor Cells with Acridine Orange^*

( Division of Biology and Growth, Sloan-Kettering Division, Cornell University Medical College, and Cytochemistry Section of Division of Pathology, Sloan-Kettering Institute for Cancer Research, New York, N.Y.)

The combined use of free-growing ascites tumor cells and purified AO has made possible quantitative cellular fluor absorption studies, as well as semiquantitative evaluation of a microfluorescence cell-viability test. The test was successful in detecting death due to acid, base, ethanol, diphenylamine diazonium bromide, aerobic autolysis, and ultrasonic radiation. However, tumor cell injury caused by x-rays, a nitrogen mustard, and bromide ion remained undetected. On the basis of these results the test is considered capable of detecting damage due to cytotoxic agents.

Green or vermilion cytoplasmic fluorescence with AO does not signify that cells are living or dead unless all the conditions of the test are met. When this is attained, red fluorescent cytoplasm is correlated with cell death. Neutral fat inclusions stained red in living and dead cells at pH 6.5 and above. Other anomalous affinitics of the stain may account for the negative test results reported by workers using bacteria and sperm (3, 6).

^* The author was aided by an Alfred P. Sloan Foundation predoctoral fellowship, and this work was supported in part by research grant C-1345 from the National Cancer Institutes of Health, Public Health Service, and by grants from the Damon Runyon Memorial Fund, the Elsa U. Pardee Foundation, and the Rippel Foundation.

Received 5/29/56.