Electron microscopic localization of acridine orange binding to DNA within rat astrocytoma C6 cells

Combination of autoradiography and ultracytochemical acridine orange method in rat astrocytoma C6 cells is described. Electron microscopic examination revealed that AO binds to DNA exclusively within the euchromatin portion of the cell nucleus. X-ray microanalysis showed that AO reaction products contain lead, uranium, and osmium. These heavy metals seem to contribute to electron density of the AO reaction products. The percentage of AO positive cells was 51.8 +/- 1.9%, whereas average labeling index of C6 cells incubated with 3H-thymidine for 30 minutes was 25.6 +/- 1.7%. About 82% of 3H-thymidine-labeled cell nuclei were AO positive and the remaining 18% were free of AO reaction products. On the other hand, about 27% of AO positive cells were not labeled with 3H-thymidine. However, most of C6 cell nuclei with active 3H-uridine incorporation showed AO chromatin reaction products. The results suggest that visualization of AO chromatin reaction products might be related to RNA synthesis rather than DNA synthesis in the cell nuclei.