Flow cytometric measurement of benzo[a]pyrene-diol-epoxide-DNA adducts in normal human peripheral lymphocytes and cultured human lung cancer cells
✍ Scribed by Reiko Shinozaki; Shoichi Inoue; Kwang-Soo Choi
- Publisher
- John Wiley and Sons
- Year
- 1998
- Tongue
- English
- Weight
- 150 KB
- Volume
- 31
- Category
- Article
- ISSN
- 0196-4763
No coin nor oath required. For personal study only.
✦ Synopsis
DNA adducts are mainly detected by 32 P-postlabeling and enzyme-linked immunosorbent assays. We have established a method for detection of benzo[a]pyrene-diol-epoxide (BPDE)-DNA adducts by flow cytometry, and have clarified the effects of the DNA adducts on cell-cycle progression and the relationship between cell-cycle phases and DNA adduct formation, using human peripheral lymphocytes and three human lung cancer cell lines. We measured the BPDE-DNA adduct levels in both lymphocytes and cancer cells by isolating nuclei, using a nuclear isolation buffer containing Triton X-100 and staining with a BPDE-DNA-specific monoclonal antibody and biotin-streptavidin fluorescein conjugates. BPDE did not affect cell-cycle progression in human peripheral lymphocytes. However, in human lung cancer cells exposed to G1 g/ml BPDE, accumulation of cells in the S phase was seen. Cells with DNA content greater than G 2 M (aneuploid cells) or cells with less than G 1 DNA content (apoptotic cells) increased gradually with exposure to increasing BPDE concentrations, suggesting that BPDE may affect cell-cycle progression through binding to DNA. Thus, the measurement of DNA adducts by flow cytometry may provide new insights into carcinogenesis.