The objectives of this study were to: (i) describe a protocol measuring the oxidation-reduction potential (ORP) by MiOXSYS System as an alternative method of seminal oxidative stress (OS) testing; (ii) establish a reference value for static ORP (sORP) to distinguish between controls and male factor infertility patients; (iii) evaluate intra-observer and inter-observer reliability; and (iv) examine association of sORP with sperm parameters predictive of male factor infertility. Elevated levels of sORP were seen in infertile patients (6.22 ± 1.10 mV/10(6) sperm/ml) compared with controls (1.59 ± 0.29 mV/10(6) sperm/ml) (P = 0.004). A sORP cut-off value 1.36 mV/10(6) sperm/ml identified normal semen and abnormal semen quality with a sensitivity 69.6%, specificity 83.1%, positive predictive value 85.3% and negative predictive value 65.9%. The test demonstrated strong intra-observer (CV 8.39%) and inter-observer reliability (correlations >0.97). Higher sORP levels were associated with poor sperm parameters across the fertility status of subjects. Negative correlations were noted with sperm parameters (concentration, total sperm count, motility and morphology) indicating these male infertility parameters are related to OS. In conclusion, the introduction of ORP as a novel clinical test for assessment of OS will help clinicians to better diagnose and manage male factor infertility patients.