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Comparison between the Comet Assay and Fast Micromethod® for Measuring DNA Damage in HeLa Cells

Nevenka Bihari ; Centre for Marine Research, Ruđer Bošković Institute, G. Paliaga 5, HR-52210 Rovinj, Croatia
Renato Batel ; Centre for Marine Research, Ruđer Bošković Institute, G. Paliaga 5, HR-52210 Rovinj, Croatia
Željko Jakšić ; Centre for Marine Research, Ruđer Bošković Institute, G. Paliaga 5, HR-52210 Rovinj, Croatia
Werner E. G. Müller ; Institute for Physiological Chemistry, Johannes Gutenberg University, Duesbergweg 6, D-55099 Mainz, Germany
Petra Waldmann ; AMMUG, Commission: Molecular Biology, Academy of Science and Literature, Obere Zahlbacher Straße 63, D-55101 Mainz, Germany
Rudolf K. Zahn ; AMMUG, Commission: Molecular Biology, Academy of Science and Literature, Obere Zahlbacher Straße 63, D-55101 Mainz, Germany


Puni tekst: engleski pdf 95 Kb

str. 793-804

preuzimanja: 524

citiraj


Sažetak

The sensitivity and precision of the single cell gel electrophoresis (Comet) assay and Fast Micromethod® for DNA damage determinations in human HeLa cell line were compared. The first assay allows analysis of DNA breaks in individual cells while the second is a rapid and convenient procedure for DNA breaks determination in cell suspensions on single microplates. Both assays detect DNA strand breaks, alkali-labile sites and transient breaks occurring at sites of ongoing repair and might be applied for the assessment of surface water genotoxic potential as well as for clinical use. DNA damage in HeLa cells was induced by different doses of γ-rays generated by Cs137 (8 to 500 cGy), UV-C light (10 to 1000 J m-2) and by different concentrations of 4-nitroquinoline-V-oxide (0.026-2.6 μmol dm-3). Gamma rays induced a dose-depended response with the average Comet tail moment values from 7 mm for the negative control to 291 mm for 200 cGy, from 6.1 to 192 mm for 500 J m-2 of UV-C light and from 7.1 to 238 mm for 1.0 μmol dm-3 of 4-nitro-quinoline-N-oxide. The Fast Micromethod® strand scission factor varied from 0.010 for negative control to 0.701 for 500 cGy, from 0.019 to 1.196 for 1000 J m-2 and from 0.003 to 0.810 for 0.5 μmol dm-3 of 4-nitroquinoline-IV-oxide. Sensitivity was the same for both methods and in the case of 4-nitroquinoline-IV-oxide even better precision (lower variation coefficient) was achieved with the Fast Micromethod®. Since the time required for multiple analysis by the Fast Micromethod® is short (2 hours or less), its use in measuring DNA breakage in cells can be recommended for environmental genotoxicity monitoring.

Ključne riječi

Fast Micromethod®, Comet assay, gamma rays, UV-C light, 4-nitroquinoline-IV-oxide, HeLa cells, DNA damage, environmental monitoring

Hrčak ID:

129322

URI

https://hrcak.srce.hr/129322

Posjeta: 932 *