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Hindi Research Article
An Efficient Technique to Detect Sperm Reactive Oxygen Species: The CellRox Deep Red® Fluorescent Probe
| Maíra Bianchi Rodrigues Alves1, André Furugen Cesar de Andrade2, Rubens Paes de Arruda3, Leonardo Batissaco1, Shirley Andrea Florez- Rodriguez1, Renata Lançoni3, Bruna Marcele Martins de Oliveira1, Mariana Andrade Torres2, Gisele Mouro Ravagnani2, Tamie Guibu de Almeida1, Vinícius Silva Vellone1 and Eneiva Carla Carvalho Celeghini1* | |
| 1Laboratory of Research in Pathology of Reproduction, Department of Animal Reproduction, School of Veterinary Medicine and Animal Science, University of Sao Paulo (USP), Pirassununga, SP, Brazil | |
| 2Laboratory of Andrology and Embryo Technology, Department of Animal Reproduction, School of Veterinary Medicine and Animal Science, University of Sao Paulo (USP),Pirassununga, SP, Brazil | |
| 3Laboratory of Semen Biotechnology and Andrology, Department of Animal Reproduction, School of Veterinary Medicine and Animal Science, University of Sao Paulo (USP), Pirassununga, SP, Brazil | |
| *Corresponding Author : | Eneiva Carla Carvalho Celeghini Animal Reproduction Department FMVZ, USP, Av. Duque de Caxias Norte 225. Postal Code 13635-900 Pirassununga/SP, Brazil Tel: +55(19) 3565-4242 E-mail: celeghin@usp.br |
| Received February 28, 2015; Accepted March 25, 2015; Published April 02, 2015 | |
| Citation: Alves MBR, de Andrade AFC, de Arruda RP, Batissaco L, Florez- Rodriguez SA, et al. (2015) An Efficient Technique to Detect Sperm Reactive Oxygen Species: The CellRox Deep Red® Fluorescent Probe. Biochem Physiol 4:157. doi:10.4172/2168-9652.1000157 | |
| Copyright: © 2015 Alves MBR, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. | |
Abstract
Heat stress and testicular traumas increase sperm Reactive Oxygen Species (ROS) resulting in Oxidative Stress (OS) and injury of sperm. The fluorescent probe CellROX Deep Red® is not already used to detect ROS in spermatozoa. On this way, this study aims to evaluate its efficiency in detecting ROS in ram sperm. For that, it was used ejaculates from rams performing the in vitro induction of sperm OS in T0, T50 and T100. T0 was semen sample that was not submitted to OS induction, T50 was 50% induced to OS induction and T100 was entire submitted to OS induction. The production of ROS was detected by CellROX Deep Red®. Data polynomial regression analysis was performed and the result of determination coefficient was 0.728. Besides, sixteen rams were submitted to scrotal insulation during 72 hours performing the in vivo induction of OS sperm. Analyses of sperm motility, morphology, DNA fragmentation and ROS (detected by CellROX Deep Red®) were done two times before the scrotal insulation and two times afterwards. Data analysis of variance was performed from the periods (before x after) and it is observed that after scrotal insulation the total and progressive motility decrease (p<0.05) and total and major sperm defects and sperm DNA fragmentation increase (p<0.05). Moreover, after insulation, it was observed increase (p<0.05) in sperm showing moderate and intense OS. Thus, it is possible to conclude that CellROX® fluorescent probe is able to detect ROS production in ram sperm by in vitro and in vivo induction being a new technique to detect sperm OS.
