Bromodeoxuridine Procedures for Analysis of DNA Synthesis
Significant advances have been made in studies on DNA synthesis, cell cycle traverse and cell proliferation through the use of radiolabeled DNA precursors. The use of tritiated thymidine to label cells actively synthesizing DNA enabled Howard and Pelc (1953) to subdivide the cell cycle in the G1, S, and G2/M phases. The capability for following the traverse of labeled S phase cells through the subsequent phases of the cell cycle has also dramatically improved investigations on cell kinetics and cell division patterns. Recently, 5-bromodeoxyuridine (BrdUrd), the base analog of thymidine, has been substituted for the radioactive precursor. The use of BrdUrd has several advantages in that (a) it can be more safely utilized in studies that would prohibit or limit the use of radiolabeled precursors, (b), methods for assaying BrdUrd, approaching the sensitivity for detection of triatiated thymidine, are currently available. One of the most widely used cytological technique for detection of BrdUrd-substituted DNA is the immunofluorescent assay developed by Grantzer (1982). This method requires partial denaturation of DNA to the single stranded conformation, by heat or acid treatment, to provide access of the incorporated BrdUrd to the antibody, which is usually tagged with the green fluorochrome, fluorescein isothiocyanate (FITC).
KeywordsPropidium Iodide Fluorescence Base Analog Thymidine Label Index Cell Division Pattern Fluorescence Difference
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