• Journal of Veterinary Clinics
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• v.17 no.2
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• pp.420-430
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• 2000

These studies were preformed to investigate the freezing conditions to achieve good post-thaw viability of spend and the practical methods of artificial insemination frozen canine semen. Semen were collected from nine male dogs which had been proved to be fertile in the past and the semen were treated for freezing procedure. Post-thaw motility and viability of canine sperm were evaluated to investigate individual tolerance of freezing, difference among freezing extenders, dif-ference among freezing equipments and freezing conditions, difference between fast and slow cooling rate, difference according to different glycerol concentration, effect of seeding on post-thaw viability, difference according to cutting part of straw, difference according to thawing temperatures, and dif-ference according to media added to thawed semen. Thawed semen for insemination were added with equal volnme of canine capacitation medium (CCM) and the volume of semen and the number per insemination were adjusted as 2-3 ml and $20-30 {\times}10^7,$ respectively. The semen were inseminated in vagina using balloon catheter and en17ryos were cellected from 9 to 11 days after the second Al to d determine fertilization.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.12
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• pp.1678-1682
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• 2001

The total proteins were estimated in both deoxycholate (DOC)-extract of sperm membrane and seminal plasma of chilled as well as frozen semen obtained from five Murrah buffalo bulls. Proteins were further characterized by polyacrylamide gel electrophoresis (PAGE) in three bulls. The protein content of sperm membrane extract (SME) and that of seminal plasma (SP) decreased gradually with increase in freezing period from 6 to 24 mo when compared with the values observed in freshly chilled semen in all bulls. The total decrease in protein content of SME and SP varied from 30-40% and 28-59% respectively during 6-24 mo of freezing. The number of glycoproteins/proteins (GP/P) in SME varied from 4-8 in freshly-chilled semen of all bulls and reduced to 2-4 after 24 mo of freezing. In SP, the number of proteins varied from 6-10 in freshly chilled semen of all bulls and reduced to 3-8 after 24 mo of freezing. Some of the proteins in SME and SP disappeared, others got altered and appeared with change in molecular weight after different freezing times. These studies reveal that alterations in the sperm membrane proteins may be responsible for damage to their membrane during freezing and thus lowering their fertilizability.

• Proceedings of the KSAR Conference
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• 2004.06a
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• pp.278-278
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• 2004

The purpose of this study was to establish the convenient freezing method for more cheap and simple. Semen quality was evaluated the motility, viability, abnormality, acrosome intactness and membrane integrity. And there were also examined the developmental rates of IVM/IVF embryos using frozen-thawed boar semen in each treatment group. (omitted)

• Animal Bioscience
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• v.36 no.10
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• pp.1530-1535
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• 2023

Objective: Semen cryopreservation result in decreased sperm parameters and fertilization ability. Sericin exhibits antioxidant activity by reducing lipid peroxidation resulting from free radicals, which can potentially improve cryopreservation outcomes. The present study aimed to examine the efficacy of various sericin concentrations supplemented with a rooster semen-freezing extender on post-thaw semen quality and fertilizing ability of sperm after cryopreservation. Methods: Semen samples were collected from 40 roosters (5 reps), then were pooled, and divided into four groups by the levels of sericin supplementation (0%, 0.25%, 0.50%, and 0.75%) in a freezing extender. Semen suspensions were loaded in medium straw (0.5 mL) and cryopreserved with the traditional liquid nitrogen vapor method. Post-thawed semen was evaluated for sperm motility, sperm viability, and lipid peroxidation. Also, the fertility test was determined. Results: The results showed that supplementation of the freezing extender with 0.50% to 0.75% sericin resulted in greater total motility and progressive motility and lower malondialdehyde levels than the other groups after cryopreservation (p<0.05). However, the viability of 0.75% decreased compared with the value of 0.50% sericin supplementation (p<0.05). Moreover, the fertility and hatchability of total eggs were significantly higher in the 0.50% sericin group than in the other groups (p<0.05). Conclusion: In conclusion, 0.50% sericin is recommended as an alternative component of the freezing extender to improve cryopreserved rooster semen.

• Journal of Veterinary Clinics
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• v.8 no.2
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• pp.183-190
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• 1991

Pellet and straw methods in canine semen freezing are compared with respect to motility, viability and acrosome demage of sperm during each of the two major processing steps, to prior-freezing and to frozen-thawing. Senen was extended with a tris-buffered egg yolk contained 4% glycero1 Pellet freezing in the hole of dry ice and straw freezing on the surface of liquid nitrogen were carried out, respectively. The frozen semen 10 days after storage in liquid nitrogen container. wao thawed. In the comparison of two freezing methods, the straw freezing method with 42.7% in motility. 49.2% in viability and 0.186 acrosome score after thawing seems to be superior to the pellet freezing method with 31.2%, 34.5% and 0.314%, respectively. Sperm motility of processing step to frozen-thawing against decrease rate 12.67% to Prior freezing appeared of 33.84% and 49.37% in straw and pellet freezing and increase of 0.02 in acrsomal score to prior freezing appeared of 0.08 and 0.21 in straw and pellet freezing method to frozen-thawing

• Journal of Embryo Transfer
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• v.29 no.4
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• pp.397-401
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• 2014

The boar sperm has more lipid droplets and specialty of seminal plasma compared with other species, causing difficulties of freezing sperm and decreases for the utilization of frozen semen into the artificial insemination. However, several studies reported significant results for the recovery of sperm motility and reproductive by addition of cryoprotectants and seminal plasma after thawing. This study was designed to investigate the effects of supplementation of trehalose or glycerol in the LEY (lactose and egg yolk in BTS) solution for the conventional freezing and vitrification process. Two boars aged 16 months were used to collect semen for 2 times in a week. The samples were allotted to 3 freezing solutions (LEY + glycerol 10.5% + OEP 1.5%, LEY + trehalose 1M + OEP 1.5%, and sucrose 1.5M + trehalose 1 M + OEP 1.5%) after centrifugation at 800 g for 10 minutes. Semen was equilibrated in freezing solutions for 10 minutes and injected into plastic straws with 2~3 air bubbles to minimize freezing damages. Vitrification was performed to locate sperm in 5 cm above $LN_2$ for 5 minutes, and the conventional freezing was conducted with an automatic freezer. Motility and survival rates were measured by CASA (Computer assisted sperm an alyzing system) and FITC (Fluorescein isothiocyanate), respectively after thawing semen at $50^{\circ}C$ for 12 seconds. The results were analyzed by ANOVA with STATVIEW statistical program. The vitrificatioin solution (LEY + 10.5% glycerol + 1.5% OEP) presented higher motility (20.9%) than other solutions while the solution (LEY + 1M trehalose + 1.5% OEP) showed the lowest (motility : 5.2%). However, survival rates of vitrified sperms detected by FITC showed 1~4% live sperms in almost of dead sperms at all vitrification solutions' groups, but survival rate of freezing solution of LEY + 1M trehalose + 1.5% OEP LEY and LEY + 10.5% glycerol + 1.5% OEP were showed 49%, and 79%, respectively. There were differences (P<0.05) survival rate of conventional freezing in LEY + 10.5% glycerol + 1.5% OEP and LEY + 1M trehalose + 1.5% OEP and the remaining showed no differences. The results suggested that vitrified boar semen was not enough to be utilized for the artificial insemination, but it showed possibility to utilize for ICSI and conventional freezing with glycerol would be useful method for artificial insemination in pig while we choose the outstanding semen against tolerance to freezing damages.

• Journal of Embryo Transfer
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• v.23 no.3
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• pp.161-166
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• 2008

The techniques for the collection, cooling and freezing of semen and artificial insemination of horses are not fully understood in Korea. We investigated the percentages of total motile (TM) and progressively motile (PM) sperms after the collection, cooling and freezing of stallion semen. The average volume of semen was 167 ml in Thoroughbred and 68 ml in Arab. The average numbers of spermatozoa in Thoroughbred and Arab were $104\times10^6/ml$ and $86\times10^6/ml$ respectively. The average percentages of TM and PM were 82.3% and 88.6% in Thoroughbred, and 61.4% and 82.6% in Arab, respectively. The average percentage of TM at 4 hr after cooling at $5^{\circ}C$ was significantly lower than that at 0 hr ($30.0\pm4.1%\;vs.\;78.0\pm2.5%,\;p<0.05$), but the percentage of PM was similar between 66.5 and 73.2% at 0, 1, and 4hr. The average percentage of frozen-thawed Thoroughbred semen frozen in MFR5 extender was 56.2%, which was significantly higher than that of the semen frozen in LE extender (average 32.9%, p<0.05). The percentage of TM in Arab was similar for semen frozen in MFR5 extender and LE extender (18.2% and 21.2%, respectively), but the percentage of PM was significantly higher in sperm frozen in MFR5 extender than in sperm frozen in LE extender (69.0% vs. 36.4%, p<0.05). Four mares were artificially inseminated by Thoroughbred frozen-thawed semen and one of them fertilized at 11 day after artificial insemination. In this study, the collection, cooling and freezing of equine semen were possible under domestic conditions.

• Journal of Veterinary Clinics
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• v.11 no.2
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• pp.545-552
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• 1994

This experiment was carried out to investigate the renditions to maintain good post-thaw motility and viability of canine spermatozoa when the semen was frozen using methanol. The semen from two male dogs which had been proven to be fertile in the previous one year was treated with different compositions of semen diluent and was frozen at different freezing temperatures, When canine semen was frozen at-2$0^{\circ}C$, -6$0^{\circ}C$, or -8$0^{\circ}C$, the spermatozoa frozen and stored at -2$0^{\circ}C$ showed very low post-thaw motility and viability from day 2 to 7 and showed no viability since day 15 after freezing. The spermatozoa frozen and stored at -6$0^{\circ}C$ or -8$0^{\circ}C$ showed higher post-thaw motility and viability on day 2, 1, 15 and 30 after freezing than that frozen and stored at-2$0^{\circ}C$(p<0.01), with no difference between two groups. Among different composition groups of the semen diluents of control(tris + egg yolk + glycerol), egg yolk-free, 히ycerol-free, and tris-free, Prior to freezing, the egg yolk-free diluent showed significantly love. motility and viability than the other diluents(p<0.05). On each thawing day (from day 2 to 15 after freezing), control group showed considerably higher motility and viability than the other groups(p<0.01). The canine spermatozoa frozen and stored at -6$0^{\circ}C$ and -8$0^{\circ}C$ showed gradual decrease of motility from day 2 to 30 after freezing and the spermatozoa of these two groups thawed on day 30 showed considerably love. motility than those thawed on day 2 after freezing, respectively(p<0.01). These results indicate that the freezing temperature of either -6$0^{\circ}C$ or -8$0^{\circ}C$ can be applicable to the freezing method using methanol and also all of the components of the semen diluent including cryoprotectant, buffer and cold-shock buffer are very important to maintain motility and viability of canine spermatozoa in the freezing and thawing procedure.

• Reproductive and Developmental Biology
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• v.30 no.1
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• pp.59-64
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• 2006

This study was conducted to establish a convenient freezing method of boar semen. Boar semen was cooled until $5^{\circ}C$ for 3 hrs using cell freezer and loaded into straws. Semen straws were frozen in different steps in strofoam box filled with $LN_2$. Highest sperm viability (54.0%) was obtained by 1-step freezing(holding at 10 cm height from the surface of $LN_2$ for 10 min). Sperm viability increased by holding at $-102^{\circ}C$ for 10min (74.0%, P<0.05). In thawing regime, sperm viability was significantly higher in $37^{\circ}C$ group than in $52^{\circ}C$ group. The sperm characteristics did not differ between 1-step and 3-step. After IVF using frozen-thawed boar semen, developmental rate of embryos to the morula+blastocyst stage was in 1-step freezing group than that of 3-step freezing group (27.5 vs 14.7%, P<0.05). The result shows that the 1-step freezing with holding at $-102^{\circ}C$ for 10min before plunging into $LN_2$ is a convenient and easy freezing method for boar semen.

• Asian-Australasian Journal of Animal Sciences
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• v.19 no.4
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• pp.486-494
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• 2006

In order to protect the spermatozoa against cold shock, hen egg yolk is widely used as a cryoprotective agent in semen freezing extenders for domestic animals. The protective action of yolk is largely presumed to be due to low density lipoproteins (LDL). The effects of LDL on sperm quality of bull and northern pike (Esox lucius) after freezing-thawing have been reported, but no study has been made to evaluate the effect of LDL on boar sperm motility and other characteristics. The experiment was carried out to investigate the effect of LDL on the freezing of boar sperm in 0.25 ml straws. The aim was to evaluate the quality of boar spermatozoa cryopreserved in the presence of LDL. Motility of semen cryopreserved in LDL was analyzed and compared to semen cryopreserved with Tris-citric acid-glucose (TCG) and Tris-citric acid-fructose (TCF), two basic freezing extenders containing egg yolk. Similarly, acrosome and plasma membrane integrity were also evaluated and compared to semen cryopreserved with TCG and TCF. Analysis of sperm quality after freeze-thaw showed that the motility, acrosome and plasma membrane integrity were improved with LDL in the extender, as compared to the TCG and TCF. The highest post-thaw integrity of acrosome and plasma membrane and motility were obtained with 9% LDL (w/v). Consequently, the optimum LDL concentration in the extender was 9%. It is also suggested that the concentration of LDL addition is important for the effect on boar sperm protection during freezing and thawing. The percentage of motile spermatozoa was significantly higher after freezing in 9% LDL than in TCG and TCF 54.4% versus 30.4% and 30.1% (p<0.05), respectively. The integrity of acrosome and plasma membrane were also significantly higher at 70.3% and 50.5% respectively with semen frozen in 9% LDL extender compared to TCG at 37.8% and 30.3% and TCF at 36.4% and 29.9%, respectively (p<0.05),. In conclusion, we propose that extender containing LDL extracted from hen egg yolk could be used as a cryoprotective media with a better efficiency than TCG and TCF. LDL improved boar semen quality, allowing better spermatozoa motility, acrosome and plasma membrane integrity after the freeze-thaw process. Furthermore, we found out that the extender with 9% LDL concentration significantly enhanced motility, acrosome and plasma membrane integrity of boar sperm after freezing and thawing.