Detection of DNA-crosslinking agents with the alkaline comet assay

✍ Scribed by Stefan Pfuhler; Hans Uwe Wolf

• Publisher: John Wiley and Sons
• Year: 1996
• Tongue: English
• Weight: 527 KB
• Volume: 27
• Category: Article
• ISSN: 0893-6692
• DOI: 10.1002/(sici)1098-2280(1996)27:3<196::aid-em4>3.0.co;2-d

✦ Synopsis

The single cell gel electrophoresis, or comet assay, under alkaline conditions is a sensitive, simple and rapid method for the detection of DNA damage at the individual cell level. Its applicability as an indicator for the DNA crosslinking potency of a test substance was investigated in human white blood cells by combined treatment with the DNA damaging agent methyl methanesulphonate (MMS) for 2 hr at 37°C. The known crosslinking agents cisplatinum, mitomycin C and formaldehyde, and the formaldehyde releasers diazolidinyl urea and dimeth-~ ~_ _ _ _ _ _ ylol urea, were shown to reduce MMS-induced DNA migration in the comet assay in a concentrationdependent manner. Two other protocols, adding MMS to the cells before or after treatment with a crosslinking agent, were carried out and achieved similar results. The results of this study indicate that the comet assay is a useful tool for the detection of crosslinking agents. Advantages and limitations of this method compared to the alkaline elution technique are discussed.