THE EFFECT OF EQUEX STM (R) IN FREEZING MEDIA ON POST THAW MOTILITY, VIABILITY AND DNA INTEGRITY OF FROZEN - THAWED RAM SPERMATOZOA

In this study we investigated the effect of Equex STM (R) on quality and in-vitro survival of ram spermatozoa frozen in Tris egg yolk based extender. Ejaculates from 6 crossbreed rams were frozen according to the standard procedure after two step dilution with Tris-egg yolk extender (1). The second extender, added to the semen at 5 degrees C, contained 14% of glycerol and was supplemented with detergent 0.75 % Equex STM (R) (group OEP) or contained no detergent (control group). After thawing the samples were incubated in a water bath at 37 degrees C and analysis were performed 10 minutes, 6, 12 and 24 hours later. Motility and the viability (Viadent (R)) of the semen were analysed with Hamilton Thorne Biosciences, Version 12.3 and membrane integrity with HOS (hypoosmotic swelling test). DNA fragmentation (DFI %) of F/T spermatozoa was analyzed 10 minutes and 3 hours after thawing using sperm chromatin structure assay (SCSA (TM)). The sperm membrane integrity was analysed 15 minutes and 3 hours after thawing by Sybr-14/PI test. Percentage of motile spermatozoa was significantly higher in OEP group in comparison to control group at 0, 6, 12 and 24h (P < 0.001). Viability of spermatozoa was significantly higher (P < 0.001) in OEP compared to control group in all analysed times after thawing. Percentage of HOS positive spermatozoa was significantly higher in OEP compared to control group respectively for 0 (P = 0.001), 6 (P =<0.001), 12 and 24h (P = 0.002) after thawing.