The aim of the study was to test commercial extenders used for short-term and long-term sperm preservation. Semen was collected in the reproduction season, i.e. from June to December. The ejaculates were obtained from single services and the routine analysis of the semen was performed immediately after the collection. The examination included semen volume, colour and texture, sperm concentration and motility, ejaculate turbulence and percentage of sperm with abnormal morphology. The semen was diluted with an extender in the ratio of 1:4. The processed semen was transported in an insulated container at 16–18 °C to the laboratory and stored in a stationary thermostat under the same temperature. Sperm motility tests were performed 24, 48, 72 and 96 hours after the placement in to thermostat. Ejaculates diluted with Ovipro, Optidyl, Triladyl and Andromed CSS gave very good results of viability (81.23 %–83.41 %) after 24 hours of storage. After 48 hours, Ovipro, Andromed, Optidyl and Triladyl gave values above 75 %. The Triladyl extender proved to be a good stabilizing agent, showing consistent results during a long-term storage. It was chosen as a control one for overall assessment. Other preservation media did not show any improving or worsening effects. The extender Ovipro showed a high motility effect in the first 48 hours only, and hence it appears to be the best solution for the short-term preservation.
The aim of this study was to compare two vitrification methods of porcine perihatching blastocysts with regard to the success of transfer of these embryos to the recipients. Expanded, hatching, or hatched blastocysts were recovered post mortem from superovulated donors in 5.5 to 6.0 days after artificial insemination of donor gilts with homospermic doses. In protocol VS I, the embryos in perihatching developmental stage were equilibrated in a culture medium H-MEMD with 10% v/v of glycerol (1.37M solution of glycerol in medium) for 10 min and placed in a vitrification medium for 1.5 min max. (vitrification medium contained 50% v/v 2M sucrose in tridistilled water, 30% v/v of glycerol, and 20% v/v of foetal calf serum – FCS). Then they were dropped with micropipette and stored in liquid nitrogen vapour. For protocol VS II, we used H-MEMD culture medium supplemented with 20% v/v of FCS, 25% v/v ethylene glycol, and 25% v/v dimethyl sulphoxide (DMSO). Embryos were equilibrated for 10 min in a mixture of the vitrification medium and culture medium (1 : 1), and were kept in the vitrification medium for 1.5 minutes. Then they were dropped with micropipette and stored in liquid nitrogen vapour. Embryos were thawed by immersing the drop with the embryo in H-MEMD culture medium with 0.8M sucrose for 10 minutes. After thawing and washing in the medium with sucrose, all embryos were washed three times in a fresh medium and prepared for transfer. Recipients were synchronized either using Regumate-feeding followed by treatment with PMSG and HCG (gilts) or using piglet weaning (sows – 1st and 2nd parity). Recipients showing standing heat at the time of donor insemination were used for laparoscopic and non-surgical ET on day 5.5–6.0 of the cycle. The fraction of viable embryo vitrified under VS I or VS II protocol was 85% and 80%, compared to 95% in control fresh embryos (P > 0.05). Pregnancy of recipients was 57.3% (5/7), 67.0% (4/6) for VS I or VS II group and 42.7% (10/23) for control (P < 0.001). We can conclude on the basis of our data that both protocols for vitrification yielded similar results and can be used for cryopreservation of porcine embryos.
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