2.5.2 The enzyme modified comet assay to detect DNA oxidation
An enzyme-modified comet assay was performed as described by Collins et al. (2023), only with peripheral blood of the mothers. For the measurement of oxidized purines, there is an additional step included after the lysis: the slides were washed three times with ice-cold buffer (0.1 M KCl, 0.5 mm Na2 EDTA, 40 mm HEPES, and 0.2 mg/ml bovine serum albumin; pH 8.0) and incubated for 40 min at 37°C with formamidopyrimidine DNA glycosylase (FPG) (Uniscience®, 1:1000) or buffer. Further steps were performed as mentioned for the comet assay under alkaline conditions (Collins et al., 2023). Net FPG sensitive sites were calculate as the difference between the samples incubated with FPG and the samples incubated with buffer; Damage Index of samplex/FPG – Damage Index of samplex/buffer.