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. Author manuscript; available in PMC: 2024 Dec 5.
Published in final edited form as: Anal Chem. 2020 Jul 8;92(14):9887–9894. doi: 10.1021/acs.analchem.0c01393

Figure 2.

Figure 2

Optimization of labeling reactions by sSingle- molecule detection and quantificationofquantification of DNA damage. HEK293 cells were exposed to DNA damage agents. The DNA samples were labeled using either the T4-PDG repair cocktail (for UVB-irradiated cells) or hOGG1 repair cocktail (for H2O2-treated cells), and the damage level was quantified.The DNA was then extracted and the damage sites were labeled with ATTO-647 fluorophores (red). The DNA molecules were stained with YOYO-1 (blue) followed by stretching on microscope cover-slip glass slides for imaging and analysis. The damage sites are seen as red dots (ATTO-647 fluorophore) on the blue (YOYO-1) DNA molecules in the representative images in A-C. (A) Control cells that were not exposed to any type of DNA damaging agent;e (B) UV-B induced DNA damage (302nm, 100 J/m2); (C) Oxidationve induced DNA damage (0.5 hours of treatment with 100 mM nM H2O2)KBrO3 for oxidative damage. (D) A plotbar graph of the calculated damage siteslevels per Mb of versus the DNA length (in base pairs). Scale bar = 20 μm.