Wayne Vessey1, Ana Perez-Miranda1, Rachel Macfarquhar1, Ashok Agarwal2, Sheryl Homa3. 1. Andrology Department, The Doctors Laboratory, London, United Kingdom. 2. Andrology Center, Cleveland Clinic, Cleveland, Ohio. 3. Andrology Department, The Doctors Laboratory, London, United Kingdom; Andrology Solutions, London, United Kingdom. Electronic address: s.homa@andrologysolutions.co.uk.
Abstract
OBJECTIVE: To standardize and validate an assay for reactive oxygen species (ROS) in human semen. DESIGN: ROS levels assayed in blanks, negative and positive control samples (30% H2O2), and human semen, with the use of a luminol-based chemiluminescence assay measured in a single tube luminometer. SETTING: Andrology laboratory. PATIENT(S): Semen samples from 19 men attending for routine semen analysis. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): ROS levels reported in relative light units (RLU) per second, adjusted for sperm concentration. RESULT(S): The ROS assay equipment performed according to expectations, generating a chemiluminescence signal for positive control samples and semen samples that decayed rapidly and was captured within 10 minutes. Blanks and negative control samples gave negligible readings. There was no significant intra- or interassay variation. Interference from extraneous factors was negligible. The assay distinguished changes in ROS over a wide range of concentrations and provided consistent results between reagent batches. Working reagents remained stable for 3 months. Acceptable levels for negative and positive control samples were established to set criteria for the test passing or failing on any given day. The assay was sensitive to ambient temperature >25°C. ROS declined significantly with time after ejaculation. Mechanical agitation doubled ROS production in semen. CONCLUSION(S): These results validate the ROS assay and demonstrate that it is a highly reliable and accurate diagnostic test.
OBJECTIVE: To standardize and validate an assay for reactive oxygen species (ROS) in human semen. DESIGN:ROS levels assayed in blanks, negative and positive control samples (30% H2O2), and human semen, with the use of a luminol-based chemiluminescence assay measured in a single tube luminometer. SETTING: Andrology laboratory. PATIENT(S): Semen samples from 19 men attending for routine semen analysis. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): ROS levels reported in relative light units (RLU) per second, adjusted for sperm concentration. RESULT(S): The ROS assay equipment performed according to expectations, generating a chemiluminescence signal for positive control samples and semen samples that decayed rapidly and was captured within 10 minutes. Blanks and negative control samples gave negligible readings. There was no significant intra- or interassay variation. Interference from extraneous factors was negligible. The assay distinguished changes in ROS over a wide range of concentrations and provided consistent results between reagent batches. Working reagents remained stable for 3 months. Acceptable levels for negative and positive control samples were established to set criteria for the test passing or failing on any given day. The assay was sensitive to ambient temperature >25°C. ROS declined significantly with time after ejaculation. Mechanical agitation doubled ROS production in semen. CONCLUSION(S): These results validate the ROS assay and demonstrate that it is a highly reliable and accurate diagnostic test.
Authors: Sheryl T Homa; Anna M Vassiliou; Jesse Stone; Aideen P Killeen; Andrew Dawkins; Jingyi Xie; Farley Gould; Jonathan W A Ramsay Journal: Genes (Basel) Date: 2019-03-19 Impact factor: 4.096
Authors: Ashok Agarwal; Neel Parekh; Manesh Kumar Panner Selvam; Ralf Henkel; Rupin Shah; Sheryl T Homa; Ranjith Ramasamy; Edmund Ko; Kelton Tremellen; Sandro Esteves; Ahmad Majzoub; Juan G Alvarez; David K Gardner; Channa N Jayasena; Jonathan W Ramsay; Chak Lam Cho; Ramadan Saleh; Denny Sakkas; James M Hotaling; Scott D Lundy; Sarah Vij; Joel Marmar; Jaime Gosalvez; Edmund Sabanegh; Hyun Jun Park; Armand Zini; Parviz Kavoussi; Sava Micic; Ryan Smith; Gian Maria Busetto; Mustafa Emre Bakırcıoğlu; Gerhard Haidl; Giancarlo Balercia; Nicolás Garrido Puchalt; Moncef Ben-Khalifa; Nicholas Tadros; Jackson Kirkman-Browne; Sergey Moskovtsev; Xuefeng Huang; Edson Borges; Daniel Franken; Natan Bar-Chama; Yoshiharu Morimoto; Kazuhisa Tomita; Vasan Satya Srini; Willem Ombelet; Elisabetta Baldi; Monica Muratori; Yasushi Yumura; Sandro La Vignera; Raghavender Kosgi; Marlon P Martinez; Donald P Evenson; Daniel Suslik Zylbersztejn; Matheus Roque; Marcello Cocuzza; Marcelo Vieira; Assaf Ben-Meir; Raoul Orvieto; Eliahu Levitas; Amir Wiser; Mohamed Arafa; Vineet Malhotra; Sijo Joseph Parekattil; Haitham Elbardisi; Luiz Carvalho; Rima Dada; Christophe Sifer; Pankaj Talwar; Ahmet Gudeloglu; Ahmed M A Mahmoud; Khaled Terras; Chadi Yazbeck; Bojanic Nebojsa; Damayanthi Durairajanayagam; Ajina Mounir; Linda G Kahn; Saradha Baskaran; Rishma Dhillon Pai; Donatella Paoli; Kristian Leisegang; Mohamed Reza Moein; Sonia Malik; Onder Yaman; Luna Samanta; Fouad Bayane; Sunil K Jindal; Muammer Kendirci; Baris Altay; Dragoljub Perovic; Avi Harlev Journal: World J Mens Health Date: 2019-05-28 Impact factor: 5.400