The role of heat shock protein in reproduction is widely known as a molecular chaperone in aiding and repairing protein
formation when stress occurred. The present objectives were to evaluate the effect of different thawing temperature and
time on the expression of HSP70 gene expression and the capacitation status in cryopreserved bovine spermatozoa. Briefly,
fresh ejaculates were obtained from three different adult bulls. The semen then underwent a sperm washing technique
known as Magnetic Activated Cell Sorting System (MACS) and later on, cryopreserved. The sperm- containing straws
were then thawed at five different thawing temperatures and time post-cryostorage; 20°C for 13 s, 37°C for 30 s, 40°C
for 7 s, 60°C for 6 s and 80°C for 5 s. The RNA was extracted from each of the sperm’s pellets and converted to cDNA
prior to the qPCR process. Capacitation status was then determined by means of CTC assay. The results showed that after
the process of amplification, there is a significant different of HSP70 gene expression in MACS process samples when the
thawing process was performed at 37°C for 30 s, with p<0.05. Furthermore, the CTC assay also showed that thawing at
the same temperature gave less capacitated spermatozoa with p<0.05. As a conclusion, MACS yield spermatozoa with a
better expression of HSP70 gene and less capacitated spermatozoa when thawing was done at 37°C for 30 s.
A novel electrophoretic separation system has been successfully applied for the preparation of human sperm prior to the execution of assisted reproductive techniques (ARTs). This new system is designed to overcome the generation of reactive oxygen species (ROS) through centrifugation in conventional sperm preparation. Since the previous study showed favorable outcomes in humans, this study intends to implement this new system for animal sperm preparation particularly in bull. Fresh semen from adult bulls were used. Optimization of the electrophoretic system for optimum bull sperm separation involved different strength of voltage and separation time. The voltages applied were 10V, 20V, 30V, 40V, 50V, and 60V. For each voltage applied, the system was operated for a duration of 12 min. An average of 10 μl fractionalized semen was taken out at the collection site at every 2-min interval. Every fractionated sperm was then evaluated for percentage of viability, motility, and DNA damage assessment. Result showed that electrophoresis at 20V and 6 min yielded more than 80% viable and more than 70% motile sperm population with the lowest DNA damage. In conclusion, the system was able to fractionate high quality bull sperm at 20V and 6 min.