Intracellular binding sites of acridine orange in living osteosarcoma cells

There have been many reports concerning the intracellular binding sites of acridine orange (AO), although the actual localization of AO in living cell s remains controversial. This study was under-taken to clarify the intracel lular localization of AO in living mouse osteosarcoma cells by cytochemical staining A mouse osteosarcoma cell line (MOS) was cultured and continuousl y et-posed to 0.5 mu g/ml of AO. The intracellular localization and stainab ility of AO the living tumor cells was morphologically detected by a high r esolution fluorescence microscope. To detect the intracellular microstructu re, cytochemical staining with rhodamin 123 for mitochondria, acid phosphat ase for lysosome, Sudan-black for fat vesicle and toluidine blue for glucos aminoglycan were performed using fixed cells. The results showed that both the nucleus and cytoplasm of tumor cells at 10 minutes after exposure to 0. 5 mu g/ml of AO emitted green fluorescence, which was especially intense in the nucleolus, bur not brilliant in the nucleus and was granular orange to red fluorescence in the perinuclear particles. This stainability of AO was different from that of rhodamin 123, Sudan-black or toluidine blue, but si milar to that of acid phosphatase. Based on these results, we conclude that the green fluorescence may have derived from AO binding to double stranded RNA,not to DNA, and that orange fluorescence may have derived from aggrega ted AO binding to lysosome.