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After secondary antibody incubation, the slices were washed with physiological saline (three 5 min washes) and then mounted onto SuperFrost Plus Slides, counterstained with Fluoromount-G medium (Southern Biotech) and coverslipped

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Future studies addressing the causal relationship between lifestyle factors and sperm epigenetic factors should ideally include longitudinal, randomised-controlled trials with the use of advanced technological tools