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Martin Nikolovski
Monika Dovenska
Dimitar Bozhinovski
Ljupcho Mickov
Branko Atanasov
Kocho Porchu
Vladimir Dzabirski
Vladimir Petkov
Toni Dovensk


The current study aimed to assess the effect of adding cryopreserving extender (soy-bean lecithin-SBLE), reduced glutathione (GSH), and seminal plasma (SP) before and after thawing on viability and velocity of cryopreserved ram sperm in liquid nitrogen. Fresh ejaculates (Ovchepolska pramenka rams, n=10) were collected and pooled. One portion was extended up to 50 million/ml with SBLE (control C-a), SBLE and GSH 5 mM (E1-a), SBLE and SP 20 vol% (E2-a), and SBLE, GSH 5 mM and SP 20 vol% (E3-a), respectively. The second portion was extended with SBLE to 100 million/ml. Both portions were cryopreserved in liquid nitrogen. Following thawing, the second portion was extended in the same manner to 50 million/ml and was separated into C-b, E1-b, E2-b, and E3-b, respectively. Each group was sampled in ten replicates immediately following thawing. Thawed samples were analyzed for viability (Hancock-2 stain), and velocity (Hamilton Thorne, USA). Each sample included at least 200 cells and the results were expressed in percent values (mean±SEM). Normality (Kolmogorov) and variance comparison (factorial-ANOVA) were performed in Statistica 8 with a significance level p<0.05. E2-a (57.58% ±2.40) and E3-a (56.94% ±1.85) yielded significantly higher viability compared to the C-a (40.73 ±1.53). There were no significant differences between C-b (50.00% ±2.33), E1-b (43.61% ±1.37), E2-b (49.16% ±1.50), and E3-b (48.50% ±1.85). In conclusion, the addition of SBLE, GSH, and SP prior vs after cryopreservation has a significant effect on thawed ram sperm viability and velocity.


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