DNA strand breaks were determined on frozen BAL cells suspension, lung (3 × 3 mm of left lobe) and liver tissue (2 × 2 mm piece of median lobe). Organ samples were snap frozen directly after dissection and kept at −80°C until analysis. Sample preparation and analysis was previously described in detail [Jackson et al., 2013 (link)]. Briefly, BAL cells, lung, or liver cell suspensions were embedded in agarose (0.7% final concentration) on TREVIGEN 20-Well CometSlides™. Slides were quickly immersed into lysing solution at 4°C and stored overnight. The next day, samples were alkaline treated and subjected to alkaline electrophoresis (pH >13) in ice cold circulating electrophoresis solution. Samples were neutralized, fixed, and later stained by SYBRGreen®. Comets were scored by the fully automated PathFinder™ system (IMSTAR, France). DNA strand breaks were quantified as % DNA in the comet tail (%TDNA) and the comet tail length (TL). The day-to-day variation and electrophoresis efficiency was validated by including PBS exposed and 60 µM H2O2-exposed A549 cells as negative and positive controls, respectively, on each slide. Control cells were exposed for 30 min at 4°C as described [Jackson et al., 2013 (link)]. The day to day variation including all slides (n = 21) from this experiment was 28%.