Abstract
| Originalsprog | Engelsk |
|---|---|
| Tidsskrift | Mutagenesis |
| Vol/bind | 23 |
| Udgave nummer | 3 |
| Sider (fra-til) | 223-31 |
| Antal sider | 8 |
| ISSN | 0267-8357 |
| DOI | |
| Status | Udgivet - 2008 |
Bibliografisk note
Keywords: Cell Nucleus; Comet Assay; DNA; DNA Damage; Data Interpretation, Statistical; Humans; Leukocytes, Mononuclear; Observer Variation; Oxidative StressAdgang til dokumentet
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Variation in assessment of oxidatively damaged DNA in mononuclear blood cells by the comet assay with visual scoring. / Forchhammer, Lykke; Bräuner, Elvira Vaclavik; Folkmann, Janne Kjaersgaard; Danielsen, Pernille Høgh; Nielsen, Claus; Jensen, Annie; Loft, Steffen; Friis, Gitte; Møller, Peter.
I: Mutagenesis, Bind 23, Nr. 3, 2008, s. 223-31.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › peer review
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TY - JOUR
T1 - Variation in assessment of oxidatively damaged DNA in mononuclear blood cells by the comet assay with visual scoring
AU - Forchhammer, Lykke
AU - Bräuner, Elvira Vaclavik
AU - Folkmann, Janne Kjaersgaard
AU - Danielsen, Pernille Høgh
AU - Nielsen, Claus
AU - Jensen, Annie
AU - Loft, Steffen
AU - Friis, Gitte
AU - Møller, Peter
N1 - Keywords: Cell Nucleus; Comet Assay; DNA; DNA Damage; Data Interpretation, Statistical; Humans; Leukocytes, Mononuclear; Observer Variation; Oxidative Stress
PY - 2008
Y1 - 2008
N2 - The comet assay is popular for assessments of genotoxicity, but the comparison of results between studies is challenging because of differences in experimental procedures and reports of DNA damage in different units. We investigated the variation of DNA damage in mononuclear blood cells (MNBCs) measured by the comet assay with focus on the variation related to alkaline unwinding and electrophoresis time, number of cells scored, as well as the putative benefits of transforming the primary end points to common units by the use of reference standards and calibration curves. Eight experienced investigators scored pre-made slides of nuclei differently, but each investigator scored constantly over time. Scoring of 200 nuclei per treatment was associated with the lowest residual variation. Alkaline unwinding for 20 or 40 min and electrophoresis for 20 or 30 min yielded different dose-response relationships of cells exposed to gamma-radiation and it was possible to reduce the variation in oxidized purines in MNBCs from humans by adjusting the level of lesions with protocol-specific calibration curves. However, there was a difference in the level of DNA damage measured by different investigators and this variation could not be reduced by use of investigator-specific calibration curves. The mean numbers of lesions per 10(6) bp in MNBCs from seven humans were 0.23 [95% confidence interval (CI): 0.14-0.33] and 0.31 (95% CI: 0.20-0.55) for strand breaks (SBs) and oxidized guanines, respectively. In conclusion, our results indicate that inter-investigator difference in scoring is a strong determinant of DNA damage levels measured by the comet assay.
AB - The comet assay is popular for assessments of genotoxicity, but the comparison of results between studies is challenging because of differences in experimental procedures and reports of DNA damage in different units. We investigated the variation of DNA damage in mononuclear blood cells (MNBCs) measured by the comet assay with focus on the variation related to alkaline unwinding and electrophoresis time, number of cells scored, as well as the putative benefits of transforming the primary end points to common units by the use of reference standards and calibration curves. Eight experienced investigators scored pre-made slides of nuclei differently, but each investigator scored constantly over time. Scoring of 200 nuclei per treatment was associated with the lowest residual variation. Alkaline unwinding for 20 or 40 min and electrophoresis for 20 or 30 min yielded different dose-response relationships of cells exposed to gamma-radiation and it was possible to reduce the variation in oxidized purines in MNBCs from humans by adjusting the level of lesions with protocol-specific calibration curves. However, there was a difference in the level of DNA damage measured by different investigators and this variation could not be reduced by use of investigator-specific calibration curves. The mean numbers of lesions per 10(6) bp in MNBCs from seven humans were 0.23 [95% confidence interval (CI): 0.14-0.33] and 0.31 (95% CI: 0.20-0.55) for strand breaks (SBs) and oxidized guanines, respectively. In conclusion, our results indicate that inter-investigator difference in scoring is a strong determinant of DNA damage levels measured by the comet assay.
U2 - 10.1093/mutage/gen006
DO - 10.1093/mutage/gen006
M3 - Journal article
C2 - 18326868
VL - 23
SP - 223
EP - 231
JO - Mutagenesis
JF - Mutagenesis
SN - 0267-8357
IS - 3
ER -