Abstract
BackgroundFreeze-drying sperm has been developed as a new preservation method where liquid nitrogen is no longer necessary. An advantage of freeze-drying sperm is that it can be stored at 4°C and transported at room temperature. Although the successful freeze-drying of sperm has been reported in a number of animals, the possibility of long-term preservation using this method has not yet been studied.Methodology/Principal FindingsOffspring were obtained from oocytes fertilized with rat epididymal sperm freeze-dried using a solution containing 10 mM Tris and 1 mM EDTA adjusted to pH 8.0. Tolerance of testicular sperm to freeze-drying was increased by pre-treatment with diamide. Offspring with normal fertility were obtained from oocytes fertilized with freeze-dried epididymal sperm stored at 4°C for 5 years.Conclusions and SignificanceSperm with –SS– cross-linking in the thiol-disulfide of their protamine were highly tolerant to freeze-drying, and the fertility of freeze-dried sperm was maintained for 5 years without deterioration. This is the first report to demonstrate the successful freeze-drying of sperm using a new and simple method for long-term preservation.
Highlights
The rat is an important animal model that has been used to help understand the etiology of many human diseases [1,2]
Sperm with –SS– cross-linking in the thiol-disulfide of their protamine were highly tolerant to freeze-drying, and the fertility of freeze-dried sperm was maintained for 5 years without deterioration. This is the first report to demonstrate the successful freeze-drying of sperm using a new and simple method for long-term preservation
The cryopreservation of rat sperm has already been reported, and offspring were obtained from oocytes fertilized with cryopreserved sperm using in vitro fertilization (IVF) [10]
Summary
The rat is an important animal model that has been used to help understand the etiology of many human diseases [1,2]. The cryopreservation of rat sperm has already been reported, and offspring were obtained from oocytes fertilized with cryopreserved sperm using in vitro fertilization (IVF) [10]. Production of offspring using IVF is efficient, a considerable degree of technical skill is required to minimize damage to sperm motility due to environmental changes such as centrifugation [11], pH, viscosity, osmotic stress [12,13], and the process of freezing and thawing [14]. Current IVF protocols are time-consuming with 5 hours incubation required for capacitation and 10 hours for penetration of sperm into oocytes in vitro [10,15]. Freeze-drying sperm has been developed as a new preservation method where liquid nitrogen is no longer necessary. The successful freeze-drying of sperm has been reported in a number of animals, the possibility of long-term preservation using this method has not yet been studied
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