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Dog sperm cryopreservation using one step dilution with glycerol-free tris extender
Md A Rahman1, S H Park1, I J Yu2
1Laboratory of Theriogenology and Reproductive Biotechnologies, College of Veterinary Medicine and Bio-Safety Research Institute, Chonbuk National University, Iksan, Republic of Korea.
Objective:
The present study was to investigate a freezing method using one step-dilution with glycerol-free TRIS extender containing 172.2 mM glucose (GFTG).
Materials And Methods:
The sperm pellet from selected ejaculates was resuspended in GFTG at 1×10(8) cells/mL. The semen was cooled for 10, 30, 50 or 70 min in GFTG at 4 degree C and was frozen in LN(2) vapor or in deep freezer (-80 degree C, DF) for 20 min before plunge into LN(2). Post-thaw sperm characteristics were examined. The phosphatidylserine (PS) translocation (Annexin V-FITC) and DNA integrity (TUNEL assay) were assessed using flow cytometry.
Results:
Progressive motility and viability were significantly higher in 50 and 70 min groups than the other groups (P<0.05). PS translocation index was significantly lower in spermatozoa cooled for 50 or 70 min compared to 10 min (P<0.05). Freezing methods using LN2 vapor showed higher progressive motility than DF method (P<0.05), while viability and DNA fragmentation were not different between two freezing methods.
Conclusion:
Cryopreservation of canine sperm cooled for 50 or 70 min following one step dilution in GFTG yields more viable sperm with lower PS translocation and freezing method using LN(2) vapor is more effective on progressive motility.
