• Asian-Australasian Journal of Animal Sciences
• /
• v.26 no.4
• /
• pp.488-500
• /
• 2013

The objective of this study was to compare the effectiveness of the protocols for superstimulation of follicular growth in Thai native heifers. Heifers (n = 20) were randomly divided into four groups of five heifers/group. Heifers were given a single dose by i.m. administration of 100 mg Follicle Stimulating Hormone dissolved in polyvinylpyrrolidone (FSHp) at 24 h. Ovum pick up (OPU) occurred at 72 h ($F_{24}O_{72}$ protocol; Group 1) or 96 h ($F_{24}O_{96}$ protocol; Group 2), and at 36 h and OPU at 72 h ($F_{36}O_{72}$ protocol; Group 3) or 96 h ($F_{36}O_{96}$ protocol; Group 4) after follicular ablation. The dynamics of ovarian follicular growth were monitored by twice-daily ultrasonographic examinations. Blood sample collections were performed every 12 h after initiation of treatment for assessment of FSH, E2 and P4 profiles. All heifers were subjected to eight repeated sequential sessions of OPU. The follicular deviation commenced $24{\pm}5.32$ h after follicular ablation in all groups. The circulatory FSH surged quickly from 24 to 36 h (>0.8 ng/ml) after follicular ablation and circulatory estrogen levels steadily increased from 36 h until OPU in all groups. At the end of the OPU sessions, the mean number of aspirated follicles/heifer/session in $F_{36}O_{72}$ protocol (Group 3) and $F_{36}O_{96}$ protocol (Group 4) were higher than in the two other groups (p<0.05). The number of cumulus-oocyte complexes (COCs), cleaved and day 8 blastocysts rates in the $F_{36}O_{72}$ protocol (Group 3) were higher than in the other groups (p<0.05). It can be concluded that a single dose i.m. administration of 100 mg FSHp at 36 h and OPU at 72 h after follicular ablation ($F_{36}O_{72}$ protocol; Group 3) was the most effective protocol for superstimulation of follicular growth for repeated OPU and subsequent in vitro embryo production in Thai native heifers.

• Journal of Embryo Transfer
• /
• v.29 no.1
• /
• pp.35-41
• /
• 2014

Laparoscopic ovum pick-up (LOPU) is a convenient method for collecting oocytes in small ruminants. LOPU has the advantage of being a less invasive means of oocyte collection, thereby allowing for a repeated usage of the oocyte donor animals. A total of 25 Korean black goats were used in the winter season (December to February) and LOPU was applied to the goats which had been treated for superovulation more than two times during the last twelve months. Estrus was synchronized with an intravaginal insert containing 0.3 g progesterone for 10 to 12 days. Ovaries were hyperstimulated with eCG 1,000 IU oneshot, FSH with eCG (50 mg / 1,000 IU; 70 mg / 500 IU; 70 mg / 1,000 IU) oneshot or FSH multiple-shot with eCG oneshot ($20mg{\times}6/300IU$) given intramuscularly 72 h prior to LOPU. For these groups, the number of follicles (mean ${\pm}$ SEM) observed which developed to larger than 2 mm in diameter were $1.6{\pm}2.5$, $4.3{\pm}3.1$, $5.5{\pm}4.2$, $6.6{\pm}2.1$ and $8.8{\pm}7.8$, respectively. Oocytes were aspirated by using OPU needles and a vacuum pump. The overall oocyte retrieval rates were 41.4%. Oocytes were matured in TCM-199 supplemented with 10% (w/v) bovine serum albumin + $10{\mu}g/ml$ FSH + $1{\mu}g/ml$ $17{\beta}$-estradiol for 27 h at $39^{\circ}C$ in 5% $CO_2$ in air. Oocytes were parthenogenetically activated by ionomycin combined with 6-diethylaminopurine (6-DMAP). Total oocyte maturation and cleavage rate were 67.3% and 78.8%, respectively. In summary, LOPU is a useful oocyte collection method in Korean black goats that can provide immature oocytes for transgenesis or nuclear transfer.

• Development and Reproduction
• /
• v.15 no.4
• /
• pp.385-391
• /
• 2011

Human in vitro fertilization and embryo transfer (IVF-ET) program is a general procedure for infertile couples since first successful delivery on 1978 in UK and Korean first on 1985. Recently in Korea, more than 42,000 cases per year of IVF-ET were performed and showed good pregnancy rates compared worldwide data. The human IVF-ET procedure use many consumables, such as ovum pick-up (OPU) needles, centrifuge tubes, culture dish, ICSI pipette, culture media and ET catheters. Major of these materials are supported by the global companies. Thus, Korean IVF-ET program might be placed unstable situation by global economical risks. These uncertain problems could be overcome by the domestic production of IVF-ET materials and consumables. Two times questionnaires for Korean clinicians and researchers about the domestic production were performed and analyzed. Many of them requested domestic OPU needles, ET catheters, culture media and ICSI pipettes under good quality control and quality assurance system. This trial may be contributed to industrialization and to global competence of Korean IVF-ET program. The results of this survey can be provide a fundamental base for development and production of domestic materials and consumables for human IVF-ET program.

• Journal of Embryo Transfer
• /
• v.25 no.1
• /
• pp.15-20
• /
• 2010

This study was performed to identify the optimal timing for oocyte donor replacement during OPU procedure. OPU was carried out to collect oocytes from every donor at an interval of $3{\sim}4$ days (2 times a week). The collected oocytes were matured in vitro in TCM-199 supplemented with 10% FBS, 10 mg/ml of FSH and 1 mg/ml of estradiol for 24 h. After 24 h of exposure to sperm, the presumptive zygotes were cultured in CR1aa medium supplemented with 4 mg/ml of BSA for 3 days before being changed to CR1aa medium with 10% of FBS for another $3{\sim}4$ days. The mean numbers of retrieved oocytes were remained constantly up to 3 months ($6.0{\pm}0.5$, $6.2{\pm}0.7$, $5.2{\pm}0.6$), but significantly decreased at over 4 to 6 months ($3.7{\pm}0.5$, $2.8{\pm}0.4$, $1.2{\pm}0.2$) (p<0.05). The blastocyst development potential was also very similar rate from 1 to 3 months (37.2%, 40.4% and 44.6%), but significantly decreased from 4 to 6 months (24.8%, 29.3% and 28.6%, respectively) (p<0.05). The production of OPU derived embryos in periods of 1 to 3 months ($2.2{\pm}0.3$, $2.5{\pm}0.3$ and $2.3{\pm}0.4$) were significantly higher than those in 4 to 6 months ($0.9{\pm}0.2$, $0.8{\pm}0.2$ and $0.3{\pm}0.2$, respectively) (p<0.05). In conclusion, the efficient periods for the production of OPU derived embryos was until 4 months, twice per week to produce over 64 transferable embryos and then replace new donor after 3 months use. The best replacement time is 3 months and could be maximized production of OPU derived embryos.

• Korean Journal of Animal Reproduction
• /
• v.24 no.1
• /
• pp.83-88
• /
• 2000

This study was conducted to determine the effect of repeated ultrasound-guided transvaginal retrieval of oocytes recovery, estrous cycle and ovarian adhesion in Korean native, Hanwoo heifers. Heifers were at unknown stages of the estrous cycle at the start of experiments in which all follicles $\geq$6mm in diameter were ablated. The results obtained in this study were as follows; Follicle developing number and oocytes collected number were no effected to repeated OPU to nine session, 4 e.a range oocytes collected to repeated OPU session. Oocytes were observated follicles were 8.7$\pm$4.2 e.a, collected oocytes were 4.1$\pm$3.4 e.a to two times collected per week and observated follicles was 10.2$\pm$6.1 e.a, collected oocytes were 4.3$\pm$2.9 e.a to one times collected per week, but no difference significantly(P＜0.05). Ovaries adhesive percentage to repeated OPU was eight ovaries adhisived(20%) of forty ovaries, three ovaries(7.5%) to 1~3 times oocytes collected, four ovaries(l0%) to 4~6 times, one ovaries(2.5%) to 7~9 times oocytes collected session. To repeated OPU effection, ovaries adhisive heifers were long estrous cycle(＞25 day) to 7 heads(87.5%) of 8 head, non-adhesive ovaries heifers were 5 heads(41.7%) were long estrous cycle to repeated OPU 12 heads. Although, now unknown about the dynamics of follicles wave and about functional changes to repeated OPU ovaries, more question about ovaries adhesive cause remain.

• Journal of Embryo Transfer
• /
• v.30 no.3
• /
• pp.149-159
• /
• 2015

This study was designed to know the possibility in repeat uses of elite donor cows for getting mass production of OPU-derived embryo production (OPU-IVP). Ultrasound transvaginal ovum pick-up (OPU) performed in 6 Korean native cows was aged 4 to 10 years old. The aspiration of immature oocytes for OPU derived embryo was carried out 2 times per week, and OPU-IVP of $1^{st}$ period was carried out 22~48 sessions from each donors. And the break time for OPU-IVP of $2^{nd}$ period after $1^{st}$ OPU from each donors were 2~25 months. The OPU-IVP of $2^{nd}$ period each donors conducted total 15~65 times for 2~8 months by an ultrasonographic, was guided follicular aspiration system. The average numbers of collected oocytes, grade 1 + grade 2(G1+G2) oocytes and cleavage embryo from $1^{st}$ period OPU-IVP were significantly differences between donors (p<0.05). Total collected oocytes of donor D were significantly higher compared with donors of A, B, C, E and F (average 17.0 per session vs. 11.2, 10.1, 8.5, 10.2 and 9.6; p<0.05) and also oocytes of G1+G2 were significantly higher compared with r A and D and subsequently to donors of B, C, E and F (average 7.9 and 8.5 per session vs. 5.0, 2.7, 6.0 and 1.6; p<0.05). Cleavage rate of donor D was significantly higher compared with donors of A, B, C, E and F (average 13.1 per session vs. 10.1, 9.1, 6.9, 8.9 and 6.7; p<0.05). The average numbers of OPU-IVP for $1^{st}$ period was significantly higher from donors of B, D and E than those from donors of A, C and F (average 6.5, 7.1 and 6.5 per session vs. 3.5, 4.2 and 2.8; p<0.05). The possibility investigation of $2^{nd}$ OPU-IVP was carried out after 2~25 months rest periods from $1^{st}$ period OPU session. Total average numbers of collected oocytes, cleavages and blastocyst development rates were significantly higher from $1^{st}$ period OPU compared with $2^{nd}$ period one (p<0.05). The OPU-IVP efficiency by break for more embryo production from elite cow was analysis comparing without rest of donor A, under 6 months rest period as B and over 6 months rest period as C and then the average numbers of collected oocytes, cleavages and blastocysts were significantly higher from A group (11.8, 9.5 and 5.2 per session) than those from B and C groups (7.9, 6.2 and 2.6 vs. 9.2, 7.5 and 3.9, p<0.05), and also C group was significantly higher than B group. In conclusion, $1^{st}$ period OPU-IVP was more efficient compared with $2^{nd}$ period repeated uses of donor, and the break times for additional production of embryo on donor were needed more than over 6 months after $1^{st}$ period OPU-IVP. This repeating uses of elite donor cows given more emphasis for getting the opportunity on mass production of elite cow OPU-IVP embryo should be increased G1+G2 possibility of genetic improvement of livestock within short period.

• Journal of Embryo Transfer
• /
• v.26 no.1
• /
• pp.19-25
• /
• 2011

This study was carried out to evaluate the effect of early pregnant cow as donor for Ovum Pick-Up (OPU) derived oocyte aspiration and embryo production in Holstein heifers. Four non-pregnant and 2 pregnant Holstein heifers were used as donor and then carried out total 17 OPU session for 10 weeks (2 times per week). Recovered cumulus-oocyte-complexes (COCs) were classified into 4 grade by oocyte cytoplasm and cumulus cells and matured in vitro in TCM-199 supplemented with 10% FBS, 10 mg/ml FSH and 1 mg/ml estradiol in 5% $CO_2$ and over 99% humidity for 24 h. After 24 h co-incubation with post-thaw sperm, the presumed zygotes were cultured in CR1aa medium with 4 mg/ml BSA for 3 days and then changed CR1aa medium with 10% of FBS for another 3~4 days. The Mean number of aspirated follicles and collected oocytes in the early stage pregnant and non-pregnant heifers were $13.0{\pm}4.3$ and $10.6{\pm}3.9$, $5.4{\pm}3.4$ and $7.7{\pm}3.6$ per session, respectively. Rate of collected oocyte from aspirated follicles were 59.2% and 50.5%, respectively. The average number of good quality oocytes (Grade I and II) in the early stage pregnant and non-pregnant heifers was $3.7{\pm}2.7$ and $4.9{\pm}2.6$ (Mean${\pm}$SD). Cleavage and blastocyst developmental rates in Grade I and II were 22.2% and 25.5%, and then $1.7{\pm}0.9$ and $1.4{\pm}1.1$ blastocyst per session, respectively. In conclusion, OPU technology can be used in early stage pregnant and non-pregnant heifers without any problem and so applied OPU derived embryo production to maximize the ability of genetically valuable females.

• Journal of Embryo Transfer
• /
• v.22 no.3
• /
• pp.191-194
• /
• 2007

The purpose of the present study was to set up basic information of size and status of ovaries by using ultrasonography to retrieve in vivo matured oocytes with ovum pick- up method. Ovaries were collected from the abattoir in Jeju in May and June which is breeding season. When the size of ovaries on ultrasonography was compared with real size measured by caliper, no significant difference was shown (p<0.05). The number of preovulatory follicles (>21mm) was investigated with ultrasonography and naked eyes. Ultrasonography group had 0.83 preovulatory follicles per ovary and naked-eye group had 0.75 preovulatory follicle per ovary and their average size was 2.86 cm and 2.34 cm, respectively. The average number of follicle was 4.25 with ultrasonography and 4.38 with naked eyes. There was no significant difference considering the size of follicle and number of follicle between ultrasonography and actual size except for the size of preovulatory follicle, suggesting that information of ultrasonography is able to use for OPU or other reproductive technology of mare.

• Journal of Embryo Transfer
• /
• v.26 no.1
• /
• pp.27-32
• /
• 2011

This study evaluated the effect of recipient's breed on the gestation length and birth weight of Hanwoo calves produced by Hanwoo ovum pick up (OPU) derived embryos. Embryos of OPU derived Hanwoo cows were transferred into the Hanwoo and Holstein recipients at 6~7 days of estrus cycle. Gestation length was expressed in days from the day of embryo transfer to the birth day of offspring, and birth weight of offspring was recorded within 24 h of birth. Breed of recipient cows (Hanwoo and Holstein) has no effect on overall gestation length ($280.9{\pm}6.2$ vs. $284.4{\pm}9.8$ days) and birth weight of calves ($23.56{\pm}3.75$ vs. $27.70{\pm}7.92\;kg$). The gestation length was higher (p<0.05) for male calves than female calves in both Hanwoo ($283.8{\pm}6.7$ vs. $277.3{\pm}3.3$ days) and Holstein ($287.3{\pm}8.9$ vs. $280.1{\pm}9.1$ days) recipient cows. Birth weight of Hanwoo calves did not differ when Hanwoo recipients ($26.5{\pm}1.3$ vs. $23.3{\pm}1.2\;kg$) were used for embryo transfer. However, male calves were heavier (p<0.05) at birth than that of female calves when embryos were transferred into the Holstein cow ($33.5{\pm}4.9$ vs. $27.8{\pm}4.9\;kg$). In conclusion, the present results demonstrated that breed of recipient cows have no effect on gestation length and birth weight of OPU derived Hanwoo calves. However, gestation length and birth weight of male calves were higher in both Hanwoo and Holstein recipient cows.

• Clinical and Experimental Reproductive Medicine
• /
• v.40 no.2
• /
• pp.90-94
• /
• 2013

Objective: To evaluate the efficacy of earlier oocyte retrieval in IVF patients with a premature LH surge on hCG day. Methods: One hundred forty IVF patients (164 cycles) with premature LH surge on hCG day were included, retrospectively. We divided them into 2 study groups: LH surge with timed ovum pick-up (OPU) 36 hours after hCG injection (group B, 129 premature cycles), and LH surge with earlier OPU within 36 hours after hCG injection (group C, 35 cycles). Control groups were tubal factor infertility without premature LH surge (group A, 143 cycles). Results: The mean age (year) was statistically higher in group C than in groups A or B ($38.2{\pm}5.4$ vs. $36.2{\pm}4.2$ vs. $36.8{\pm}4.9$, respectively; p=0.012). The serum LH levels (mIU/mL) on hCG day were significantly higher in group B and C than in group A ($22.7{\pm}14.9$ vs. $30.3{\pm}15.9$ vs. $3.2{\pm}2.9$, respectively; p>0.001). Among groups A, B, and C, 4.9%, 31.7%, and 51.4% of the cycles, respectively, had no oocytes, and the overall rates of cycle cancellation (OPU cancellation, no oocyte, or no embryos transferrable) were 15.4%, 65.9%, and 74.3%, respectively. The fertilization rate (%) was significantly higher in group B than in group C ($73.2{\pm}38.9$ vs. $47.8{\pm}42.9$, p=0.024). The clinical pregnancy rate was significantly higher in group C than in groups A and B (44.4% vs. 27.3% vs. 9.1%, respectively, p=0.021). However, the miscarriage rate was also higher in group C than in group B (22% vs. 0%, respectively, p=0.026). Conclusion: Earlier OPU may not be effective in reducing the risk of cycle cancellation in patients with premature LH surge on hCG day. A larger scale study will be required to reveal the effectiveness of earlier ovum retrieval with premature LH surge.