High Variability in Human Sperm Membrane Potential Over Time Limits Its Reliability as a Predictor in ART Outcomes

Sperm membrane potential (Em) hyperpolarization during capacitation is a functional hallmark of fertilizing ability and has been proposed as a predictive biomarker for conventional in vitro fertilization (IVF) success. However, it is unclear whether Em remains stable across ejaculates over time and can reliably guide assisted reproductive treatment (ART) decisions in advance. Thus, we aimed to evaluate the temporal consistency of human sperm Em within individuals and assess its utility as a prognostic marker when measured days or weeks prior to IVF procedures. Em was assessed in capacitated and non-capacitated sperm from normospermic donors at three time points over 28 days, using a fluorometric assay. Capacitated values were compared to a −48.6 mV threshold previously associated with successful fertilization. Intra-donor Em variability and coefficients of variation (CV) were analyzed statistically. Our results showed that Em values exhibited significant intra-donor variability over time (p=0.007), with approximately half of the donors crossing the −48.6 mV functional threshold across sessions. Capacitated sperm samples showed significantly greater variability than non-capacitated ones, with several donors exceeding a 30% CV cutoff. No consistent correlation was found between CV and mean Em values. While Em remains a promising functional marker when assessed on the day of IVF, its temporal variability undermines its reliability as a predictive tool for ART decisions made in advance. These findings underscore the importance of timing in functional sperm assessments and call for further studies to identify the physiological factors influencing Em stability.