FIG. 4.

Inducible expression of AML1-ETO leads to apoptosis in U937T-A/E cells. Detection of apoptotic cells was performed by acridine orange and ethidium bromide double staining. U937T-A/E cells were maintained for 72 h in the medium either containing 1 μg of tetracycline/ml (A) or not containing tetracycline (B). Both live and dead cells take up acridine orange. Acridine orange intercalates into DNA and RNA, making the former appear green while the latter stains red. Thus a viable cell has bright green chromatin in its nucleus and red-orange cytoplasm. Ethidium bromide is only taken up by nonviable cells. Ethidium bromide intercalates into DNA, making it appear orange, but binds only weakly to RNA, which may appear slightly red. Thus a dead cell has bright orange chromatin (the ethidium overwhelms the acridine) and its cytoplasm, if it has any contents remaining, appears dark red. Cells that have undergone necrosis have the fluorescent features of nonviable cells but do not have apoptotic nuclear morphology. VN, viable cells with normal nuclei (bright green chromatin with organized structure); VA, viable cells with apoptotic nuclei (bright green chromatin which is highly condensed or fragmented); NVA, nonviable cells with apoptotic nuclei (bright orange chromatin which is highly condensed or fragmented).