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. 2019 Nov 25;9:17478. doi: 10.1038/s41598-019-53983-9

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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RF-EME exposure induces oxidative DNA damage in spermatozoa. Spermatozoa were isolated from the cauda epididymis of untreated control animals (UT), as well as those of the sham and RF-EME exposure groups. These cells were assessed for DNA fragmentation using (a) the halo assay showing the percentage of cells fragmented (n = 5–8 mice/treatment group, each with 100 sperm assessed for each replicate) and then (b) quantified by the alkaline comet assay, expressed as percentage tail intensity and normalized to control data for each run (n = 3 mice/treatment, each with 50–70 sperm cells assessed). (c) To extend this DNA integrity analysis, sperm were evaluated for oxidative DNA adducts via labelling with anti-8-hydroxy-2-deoxyguanosine (8-OH-dG) antibodies (n = 3–5 mice/treatment). (d) Representative images of spermatozoa stained with the 8-OH-dG antibody from the 5 week sham and RF-EME exposed populations are included. The number of biological replicates used is denoted in each bar. Data are presented as mean + SEM. *P < 0.05, **P < 0.01.