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. 2019 May 16;9:7492. doi: 10.1038/s41598-019-43981-2

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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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Variation of sDF during selection as evaluated by traditional techniques (revealing total sDF) and liveTUNEL (revealing viable sDF). LiveTUNEL unmasks samples increasing sDF respect to traditional techniques. (A) A sample with an increase of viable sDF (pink arrow) which is higher than the decrease due to deletion of DNA fragmented dead spermatozoa (blue arrow). In this case, both traditional techniques (total sDF, green arrow) and LiveTUNEL (viable sDF, pink arrow) can identify the increasing sample. (B) A sample with an increase of viable sDF (pink arrow) which is lower than the decrease due to deletion of DNA fragmented dead spermatozoa (blue arrow). In this case only LiveTUNEL (viable sDF, pink arrow) can identify the increasing sample. Note also that traditional techniques reveal only a part of increase (closed green arrow in A). Conversely, LiveTUNEL (pink arrow) detects all the increase. Dashed lines represent the amount of increase of sDF not detected by traditional techniques.