Variation in the measurement of DNA damage by comet assay measured by the ECVAG{dagger} inter-laboratory validation trial

Research output: Contribution to journalJournal articleResearchpeer-review

  • Lykke Forchhammer
  • Clara Johansson
  • Lennart Möller
  • Roger W L Godschalk
  • Sabine A S Langie
  • George D D Jones
  • Rachel W L Kwok
  • Andrew R Collins
  • Amaya Azqueta
  • David H Phillips
  • Osman Sozeri
  • Maciej Stepnik
  • Jadwiga Palus
  • Ulla Vogel
  • Håkan Wallin
  • Michael N Routledge
  • Catherine Handforth
  • Alessandra Allione
  • Giuseppe Matullo
  • João Paulo Teixeira
  • Solange Costa
  • Patrizia Riso
  • Marisa Porrini
The comet assay has become a popular method for the assessment of DNA damage in biomonitoring studies and genetic toxicology. However, few studies have addressed the issue of the noted inter-laboratory variability of DNA damage measured by the comet assay. In this study, 12 laboratories analysed the level of DNA damage in monocyte-derived THP-1 cells by either visual classification or computer-aided image analysis of pre-made slides, coded cryopreserved samples of cells and reference standard cells (calibration curve samples). The reference standard samples were irradiated with ionizing radiation (0-10 Gy) and used to construct a calibration curve to calculate the number of lesions per 10(6) base pair. All laboratories detected dose-response relationships in the coded samples irradiated with ionizing radiation (1.5-7 Gy), but there were overt differences in the level of DNA damage reported by the different laboratories as evidenced by an inter-laboratory coefficient of variation (CV) of 47%. Adjustment of the primary comet assay end points by a calibration curve prepared in each laboratory reduced the CV to 28%, a statistically significant reduction (P < 0.05, Levene's test). A large fraction of the inter-laboratory variation originated from differences in image analysis, whereas the intra-laboratory variation was considerably smaller than the variation between laboratories. In summary, adjustment of primary comet assay results by reference standards reduces inter-laboratory variation in the level of DNA damage measured by the alkaline version of the comet assay.
Original languageEnglish
Issue number2
Pages (from-to)113-23
Number of pages10
Publication statusPublished - 2010

ID: 18360127