Non-invasive chemical assessment of living human spermatozoa

Confocal Raman microspectroscopy was used to chemically image single, living and immobilized human spermatozoa under near-physiological conditions. A novel combination of sperm preparation, substrate and coating was used. We have investigated systematically the exposure of sperm cells to Raman laser light with varying radiation intensity. Using fluorescence microscopy we could show that at 30 mW laser power sperm cell membranes suffer from radiation damage. When illuminating with 15 mW laser power or less we observed no cell membrane damage. Several single spermatozoa have been measured in an automated fashion resulting in full chemical images including reference background with one cell per 60 s. During automated measurements the laser illumination time was decreased by 60% compared to the parameters that proved to yield no damage for safety reasons. Our novel assay is able to provide chemical information of single, live human spermatozoa in near-physiological conditions even during repeated optical measurements.