Analysis of bromodeoxyuridine incorporation into DNA: Comparison of gas chromatographic/mass spectrometric, CsCl gradient sedimentation, and specific radioactivity methods
✍ Scribed by Jonathan Maybaum; Matthew G. Kott; Norma J. Johnson; William D. Ensminger; Philip L. Stetson
- Publisher
- Elsevier Science
- Year
- 1987
- Tongue
- English
- Weight
- 688 KB
- Volume
- 161
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
A sensitive new method for the quantitation of 5-bromodeoxyuridine (BrdUrd) incorporated into DNA by CC/MS analysis of enzymatically released Thy and bromouracil (BrUra) is presented. The hydrolysis procedure was characterized and found to give uniform results when sample size was 1-10 rug DNA and incubation time for DNA digestion was between 40 min and 16 h. Samples of DNA containing 'H-labeled BrdUrd were analyzed in parallel by the GC/MS technique and by specific radioactivity and buoyant density measurements, in order to compare the three methods. The GC/MS procedure gave values for percentage replacement of Thy by BrUra which were higher than those obtained by specific activity and lower than those obtained by buoyant density. This GC/MS method can detect 1% replacement in a I-pg DNA sample, equivalent to approximately 10s cells or 0.1 mg tissue, and will permit sensitive and quantitative analysis of the presence of this chemotherapeutic agent/radiosensitizer in cellular DNA from biopsy samples of normal or tUmOr tissue.