Evaluation of rat sperm by flow cytometry: simultaneous analysis of sperm count and sperm viability

In this study, we conducted a simultaneous analysis of sperm count and viability in rats by flow cytometry (FCM). Epididymal fluids were taken from the caudal epididymis of 12 to 13 week-old Sprague-Dawley rats. The fluids were weighed and mixed with Dulbecco's phosphate buffered saline (D-PBS). Propidium iodide, which can stain only dead sperm, was used to distinguish viable and dead sperm. The sperm count and viability analyzed by FCM were 1.28 x 10(6)/mg and 78.0%, respectively. These values were consistent with the corresponding values (1.39 x 10(6)/mg and 81.0%) that were directly determined microscopically in the fluids of the same sample. In addition, when the original mixture containing sperm was diluted two times and four times with D-PBS, or was diluted two times with D-PBS containing only killed sperm, the sperm count and viability determined by FCM also correlated well with the sperm count (r = 0.96, P < 0.01) and sperm motility (r = 0.99, P < 0.01) by direct microscopic observation, respectively. In conclusion, the present flow cytometric analysis would be practical for the simultaneous determination of sperm count and viability in rat epididymal fluids.