Immunocytochemistry of apoptosis induced by bromodeoxyuridine in human leukemic HL-60 cells

Various antitumor drugs which target tumor cells in the S-phase of the cell cycle induce apoptosis in HL-60 cells. The present study shows that apoptosis is inducible in HL-60 cells by continuous exposure to bromodeoxyuridine (BrdU), an S-phase specific reagent and a thymidine analogue. The localization of BrdU in the apoptotic cells was visualized with immunogold particles using an anti-BrdU monoclonal antibody. After 3 days of treatment with 20 microM BrdU, cells ceased to grow and their viability was significantly decreased. Morphologically, nuclear segmentation and cytoplasmic maturation were observed in many viable cells, and the dead cells showed typical features of apoptosis. The nuclear DNA of apoptotic cells was visualized by the immunocytochemical detection of BrdU with post-embedding immunogold staining. The condensed chromatin of the apoptotic cells was highly labeled with immunogold particles, while the nucleolus was sparsely labeled. The chromatin could be traced with BrdU-immunocytochemistry even after disappearance of the nuclear envelope. These results indicated that apoptosis in HL-60 cells was induced by incorporation of BrdU into DNA of the S-phase cells. Using BrdU-immunocytochemistry, the localization of nuclear DNA in apoptotic cells was visualized and the changes of nuclear structures were followed during the progression of apoptosis.