Please use this identifier to cite or link to this item: http://hdl.handle.net/1942/33016
Title: An optimized comet-based in vitro DNA repair assay to assess base and nucleotide excision repair activity
Authors: Vodenkova, Sona
Azqueta, Amaya
Collins, Andrew
Dusinska, Maria
Gaivao, Isabel
Moller, Peter
Opattova, Alena
Vodicka, Pavel
Godschalk, Roger W. L.
LANGIE, Sabine 
Issue Date: 2020
Publisher: NATURE RESEARCH
Source: NATURE PROTOCOLS, 15 (12) , p. 3844 -3878
Abstract: This optimized protocol (including links to instruction videos) describes a comet-based in vitro DNA repair assay that is relatively simple, versatile, and inexpensive, enabling the detection of base and nucleotide excision repair activity. Protein extracts from samples are incubated with agarose-embedded substrate nucleoids ('naked' supercoiled DNA) containing specifically induced DNA lesions (e.g., resulting from oxidation, UVC radiation or benzo[a]pyrene-diol epoxide treatment). DNA incisions produced during the incubation reaction are quantified as strand breaks after electrophoresis, reflecting the extract's incision activity. The method has been applied in cell culture model systems, human biomonitoring and clinical investigations, and animal studies, using isolated blood cells and various solid tissues. Once extracts and substrates are prepared, the assay can be completed within 2 d. This protocol describes a comet-based in vitro assay for detecting base and nucleotide excision repair activity for use in cell culture model systems, human biomonitoring and clinical investigations, and animal studies, using isolated blood cells and various solid tissues.
Notes: Langie, SAS (corresponding author), Maastricht Univ, Sch Nutr & Translat Res Metab NUTRIM, Dept Pharmacol & Toxicol, Maastricht, Netherlands.; Langie, SAS (corresponding author), Hasselt Univ, Ctr Environm Sci, Hasselt, Belgium.
s.langie@maastrichtuniversity.nl
Other: Langie, SAS (corresponding author), Maastricht Univ, Sch Nutr & Translat Res Metab NUTRIM, Dept Pharmacol & Toxicol, Maastricht, Netherlands ; Hasselt Univ, Ctr Environm Sci, Hasselt, Belgium. s.langie@maastrichtuniversity.nl
Keywords: Oxidatively Damaged Dna;Cancer Patients;Polyphenolic Compounds;Occupational-Exposure;Incision Activity;Expression Levels;Telomere Length;Mineral Fibers;Cross-Links;Cells
Document URI: http://hdl.handle.net/1942/33016
ISSN: 1754-2189
e-ISSN: 1750-2799
DOI: 10.1038/s41596-020-0401-x
ISI #: WOS:000590016600002
Rights: 2020 Springer Nature Limited.
Category: A1
Type: Journal Contribution
Validations: ecoom 2021
Appears in Collections:Research publications

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