A new method for evaluating the quality of single sperm by detecting reactive oxygen species

Abstract

Sperm damage caused by reactive oxygen species (ROS) is one of the main causes of male infertility. Therefore, the level of ROS in sperm is an important indicator for the diagnosis and prognosis of male infertility. Herein, we constructed a single sperm ROS detection method (SSRDM) with an optical micro-probe fabricated via focused ion beam process. The micro-probe is used to separately excite fluorescence in the sperm and the area around the sperm after ROS staining, and the difference in fluorescence values can reflect the level of ROS in the sperm. We collected 102 semen samples and 72 of them were divided into asthenozoospermia and non-asthenozoospermia groups. SSRDM and flow cytometry were used to detect the ROS levels of the two groups. The results of SSRDM showed that the ROS levels of asthenozoospermia group were higher than that of non-asthenozoospermia group (P = 0.002), while the results of flow cytometry indicated no difference (P = 0.152). In terms of ROS levels, compared with flow cytometry, SSRDM has a stronger ability to distinguish between those two groups, providing a reliable basis for assessment of sperm quality. Another 30 semen samples were used to investigate temperature and temporal variability of SSRDM to ensure the stability and accuracy of this method. Overall, we have developed a method that can quantitatively detect fluorescent substances in sperm at the single-cell level supplying evidence for diagnosis and prognosis of male infertility.