Surface changes associated with ram sperm cryopreservation revealed by counter-current distribution in an aqueous two-phase system. Effect of different cryoprotectants.
Ram sperm was frozen in the presence of the most commonly used cryoprotectants. After thawing, the overall cell surface changes provoked by freezing were assessed by centrifugal counter-current distribution (CCCD). In addition, cell membrane integrity (viability) of all the treated sperm was estimated by fluorescent staining. Fresh and refrigerated sperm were used as controls. Our results show no improvement of the cooling-induced cell surface damage by freezing in the presence of bovine seminal plasma, proline, glycine-betaine and phosphatidylcholine. Better results were obtained with vitamin E and cholesterol. However, the best protective effects were found by employing seroalbumin and lactalbumin. Furthermore, freezing in the presence of bovine lactalbumin resulted in a good maintenance of the cellular viability and of the CCCD heterogeneity in respect to fresh cells.