Rapid staining methods for analysis of deoxyribonucleic acid and protein in mammalian cells

Abstract

Quantitative fluorescent staining and analysis of cellular DNA were accomplished using 3 groups of reagents having different mechanisms of action for DNA binding. These reagents included the fluorescent antitumor antibiotics mithramycin, chromomycin A3 and olivomycin; the Feulgen reagents acriflavine and flavophosphine N, and the intercalating dyes ethidium bromide and propidium iodide. Propidium iodide (PI) was used in combination with fluorescein isothiocyanate (FITC) to stain both cellular DNA and protein, respectively. Multiparameter analysis of PI/FITC stained cells provided a direct correlation of DNA and protein for cells in all stages of the cell cycle. Nuclear to cytoplasmic ratio determinations were also performed on PI/FITC stained cells by analysis of the time duration of the red (DNA) and green (protein) fluorescence signals from each cell. These staining and analysis techniques provide alternative methods for directly determining the quantitative relations between cellular DNA and protein and will be extremely useful in investigations where fluctuations of these parameters are of importance for assessing the results.