An improved DAPI-DNA cytofluorometry by hypotonic cytolysis. Its application to tumor cell heterogeneity.
The effect of cell cytolysis on DAPI-DNA cytofluorometry was studied in order to eliminate nonspecific cytoplasmic hindrances in DAPI staining. A great improvement in the reproducibility of DNA determination was attained when cells were treated with hypotonic KCl (0.075 M) solution. The DNA content per cell of the mgC5 clonal line (normal diploid cell containing 2C DNA units) after hypotonic treatment was 3.07 +/- 0.06, while that of mgC5 cells without hypotonic treatment was 3.50 +/- 0.26, indicating a remarkable decrease from 7.4% to 2.0% in the coefficient of variation. In detecting tumor cell heterogeneity, our cytolytic method was superior to the method of counting the mode of chromosome number. The modal chromosome numbers of nontumorigenic m and its derivatives, mgC4 and mgC5, were 69, 68 and 67 respectively, while those of their tumorigenic counterparts, magc, mgC4V1 and mgC5V1 were 68, 66 and 67. The DNA contents of nontumorigenic m, mgC4 and mgC5 were 3.00 +/- 0.10 (mean +/- SD), 3.07 +/- 0.06 and 3.07 +/- 0.12, while those of their tumorigenic counterparts, magc, mgC4V1 and mgC5V1 were 3.67 +/- 0.15, 3.23 +/- 0.06, 3.27 +/- 0.06 respectively. A statistically significant difference in DNA content between the tumorigenic and nontumorigenic cells was observed at least at the p less than 0.05 level (t-test) when DNA content was employed for comparison, but not when the modal chromosome number was employed.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D002478:Cells, Cultured; D002875:Chromosomes; D004273:DNA, Neoplasm / Q000032:analysis*; D005434:Flow Cytometry; D007038:Hypotonic Solutions; D007211:Indoles; D051379:Mice; D009374:Neoplasms, Experimental / Q000032:analysis / Q000473:pathology*
D004273:DNA, Neoplasm; D007038:Hypotonic Solutions; D007211:Indoles; C007293:DAPI