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Abstract
The present work was undertaken to determine the effect of cryopreservation on viability, integrity of sperm DNA and fertility of buffalo spermatozoa as well as minimizing hazards effect of freezing procedures by adding certain additives to buffalo semen.
Three mature buffalo bulls aged 4-6 years old were used in the present work. They were raised on the farm of Animal Reproduction Research Institute (ARRI, Al-Haram, Giza). Semen was collected twice a week using an artificial vagina. Two successive ejaculates were regularly collected at early morning. Immediately after collection, semen samples were evaluated. The samples that had more than 70% progressive motility and sperm concentration 6x 106 /ml were used.
Two trials were conducted in the current study. The trial 1 was carried out to detect the effect of different additives on the motility and morphology of frozen buffalo semen. Pooled semen samples were split immediately after collection into five portions and diluted 1:5 at room temperature with optidyl extender supplemented with.
1- 1- cyclodextrin (800.00 mg/mi).
2- Selenium ACE. (Selenium 250 mg/lOOml) + Vit. A
(500 IU/lOOml) +Vit.C (300 mg/lOOml) +Vit. E (100
mg/i OOml).
4- palmitic acid (100mg/i OOml).
5- Control (no additives).
The extended semen was stored at 5°C for 72 hours. Sperm motility was subjectively assessed immediately after dilution as well as after 24, 48 and 72 hours of incubation period at 5°C. The viability index (VI) was calculated.
IN Frozen storage:
Semen samples were diluted as previously mentioned in chilled storage, then the diluted semen was cooled at 5°C over a period of 45 to 60 minutes in cold-handling cabinet. The cooled semen was loaded into 0.25 ml PVC ministraws (IMV, L, Aigle, France). The straws were then arranged horizontally on the freezing racks at 5°C and were suspended in liquid nitrogen vapor inside a foam box. The frozen straws were stored in the liquid nitrogen for two weeks before thawing.
Trial II:
Individual semen samples were collected from each buffalo
bull and divided into five portions. They were diluted at rate of 1:5 with “Optidyl” extender as mentioned before in Trial I.
The freezing- thawing process and post thaw evaluation were done as previously mentioned before.
Sperm morph ologv:
Sperm morphologic abnormalities were assessed immediately after thawing by using spermac stain.