Some quantitative aspects of Acridine Orange fluorescence in unfixed, sucrose-isolated mammalian nuclei

Nuclei were isolated from mouse liver and central nervous system (CNS). These nuclei were fluorochromed without fixation in a 0.25 M sucrose medium containing 5-5 X 10(-5) M Acridin Orange and measured with an incident microfluorometric system. In the case of mouse CNS nuclei, the major and minor axes of the nuclei were measured with a filar micrometer. Three modal values were obtained from the hepatocyte suspension corresponding to 2C, 4C, and 8C nuclei, respectively. While the CNS nuclei displayed substantial variability in size, the Acridine Orange emission values at 530 nm were nearly constant. The data suggest that under these conditions, Acridine Orange fuorescence reflects DNA content. Further, the 530 nm fluorescence emission is not affected by chromatin condensation or proteins complexed with DNA.