الفهرس 
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Abstract
The present work was designed to determine the effect of relaxin
hormone and methyl-β-cyclodextrin (MBCD) on viability, motility, capacitation and fertilizing capability of buffalo frozen/thawed semen.
Spermatozoa were prepared from frozen/thawed buffalo semen, obtained
from bulls previously tested for IVF.
Sperm viability was assessed by Trypan blue/Giemsa staining technique. Sperm capacitation rate was assessed by using several techniques including Trypan blue/Giemsa staining technique, Chlortetracycline (CTC) staining technique and immune-localization of tyrosine phosphorylated protein.
The sperm fertilizing capacity was determined by assessing the penetration, cleavage, normal fertilization and polyspermic rates after heterologous IVF by staining the embryos with DAPI for nuclei examination under Epi-Fluorescence microscopy after zona removal by
pronase (2 mg/ml) digestion.
a) Effect of relaxin hormone
The effect of 3 different concentrations of relaxin (25, 50 and
ng/ml) on sperm was tested in 2 experiments; Experiment 1:
Effect of different concentrations of Relaxin hormone (25, 50 and 100 ng/ml) on livability, Motility, capacitation and Acrosome reaction (AR) of buffalo frozen/thawed semen. - Semen was divided into 5 groups: non capacitating medium (NCM, without capacitating agent), Capacitating medium (CM, contain 0.01 mM heparin) and 3 Relaxin treated groups containing 25, 50 and 100
ng/ml of Relaxin, and incubated for 2 different incubation times; 2 and
4hs in a controlled gas atmosphere of 5% CO2 in humidified air.- Following each incubation period, sperm motility was examined under the phase contrast microscope and capacitation and acrosome reaction were assessed as mentioned before. - The immune-localization of tyrosine phosphorylated protein was made only in the 4 experimental groups; NCM, CM, and the 2 Relaxin groups (50 and 100 ng/ml) as the results of trypan blue/Giemsa technique indicated that the highest values of capacitation was obtained with the concentrations 50 and 100 ng/ml Relaxin. - Statistical analysis revealed that: 1- Sperm livability was not affected by any of the relaxin treatments after 2 and 4 hs of incubation and remained high in all groups after the different incubation periods (98.22±0.54 - 98.94±0.33%).
2- Relaxin improves sperm motility after 2 and 4 hs of incubation with
concentrations of 50 (67.8±1.0, 63.9±1.1%, respectively) and 100
(67.2±1.7, 63.9±1.1%, respectively) ng/ml compared to the NCM(63.3±1.1, 56.1±1.8%, respectively) and CM (60.0±1.1, 53.3±1.1%, respectively).