• Title, Summary, Keyword: in vitro culture

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Studies on the Survival and In Vitro Developmental Rates after Bisection of Bovine Embryos (소 초기배 분할후 생존성과 체외발생율에 관한 연구)

  • ;Y. Noriko
    • Korean Journal of Animal Reproduction
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    • v.21 no.3
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    • pp.275-280
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    • 1997
  • This study was carried out to investigate on the survival and in vitro developmental rates of bisected bovine embryos by microblade, micropipette and pronase methods. Bisected embryos cultured for 1∼7 days in TCM-199 media with 10 FCS+hormones. Survival and in vitro developmental rates was defined on in vitro culture or FDA-test. The results are summarized as follows ; 1. The survival and in vitro developmental rates of bisected bovine embryos by microblade, micropipette and pronase methods were 22.2, 16.7, 15.0% and 22.2, 23.3, 18.8%, respectively. In vitro developmental rate of bisected bovine embryos was significantly lower than that of non-bisection embryos(27.8% and 25.0%). 2. In vitro developmental rates of bovine embryos bisected for 1, 2, 4, 8, 16 cells stages during in vitro culture in 10% FCS+TCM-199 media were 25.0, 20.0, 20.0, 15.0 and 6.7%, respectively. 3. In vitro developmental rates of intact and free-zona pellucida of bisected demi-embryos during in vitro culture in 10% FCS+TCM-199 media were 25.6, 16.7%, respectively. 4. In vitro developmental rates of biopsied embryos and biopsied blastomeres during in vitro culture in 10% FCS+TCM-199 media were 20.0, 11.1%, respectively.

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In Vitro Fertilization and Development of Bovine Oocytes (우 난포란의 체외수정과 발육)

  • 김정익
    • Korean Journal of Animal Reproduction
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    • v.13 no.2
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    • pp.98-104
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    • 1989
  • Successful techniques of in vitro fertilization(IVF) are valuable for studying the process of fertilization and for developing economical procedures for gene and nuclear transfer in farm animals. To date, bovine IVF system has been developed with oocytes in vitro or vitro, but the resulting zygotes exhibit limited embryonic development after in vitro culture. Even though in vitro matured oocytes achieved high fertilization and cleavage rates, these embryos appear extremly low rate of pregnancies when transferred to synchronized recipients. Development of early bovine embryos in vitro is generally arrested at the 8-to 16-cell stage. However, recent use of somatic cells such as trophoblastic vesicle, granulosa and oviduct epithelial cell for co-culture with early bovine embryos has proven effective for development of embryos, matured and fertilized in vitro, past the in vitro cell blocks. These factors clearly indicate the value of the co-culture system in promoting development of bovine oocytes matured and fertilized in vitro to morula or blastocyst stage in vitro. In addition, co-culture system may beome a tool for evaluation of viability of ova that have been manipulated by procedures such as splitting, microinjection and nuclear transfer.

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Viability of In Vitro Fertilized Bovine Embryos Following In Vitro Culture and Embryo Transfer (소 체외수정란의 체외배양 및 이식후 생존성)

  • 정희태;유재원;박연수;양부근;김정익
    • Journal of Embryo Transfer
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    • v.9 no.3
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    • pp.221-227
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    • 1994
  • This study was conducted to examine the condition of in vitro culture system and the viability after embryo transfer of in vitro matured-in vitro fertilized (IVM-IVF) bovine embryos. The in vitro development to the blastocyst stage was enhanced by supplying bovine serum albumin(BSA) to co-culture medium with bovine oviduct epithelial tissue(BOET) compared with that in medium supplemented with fetal bovine serum(FBS) (41.2% vs. 26. 3%, P<0.05). After transfer of IVM-IVF blastocysts into the uterine horn of recipient females (Aberdeen Angus), one was pregnant to term and produced a head of male Korean native calf. These results confirm that the in vitro development of IVM-IVF bovine embryos is affected with different protein source in co-culture with BOET, and IVM-IVF embryos can develop to term after in vitro culture and embryo transfer.

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Effect of MEM Vitamins Supplementation of In vitro Maturation Medium and In vitro Culture Medium on the Development of Porcine Embryos

  • Kim, J.Y.;Lee, E.J.;Park, J.M.;Park, H.D.
    • Asian-Australasian Journal of Animal Sciences
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    • v.24 no.11
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    • pp.1541-1546
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    • 2011
  • This study was carried out to examine the influence of minimum essential medium (MEM) vitamins supplementation to in vitro maturation medium and in vitro culture medium on the development of porcine embryos. Porcine embryo development was investigated following cultivation in both in vitro maturation and culture medium with the supplementation of MEM vitamins (0, 0.1, 0.2 and 0.4%) using immature oocytes collected from the ovary of prepubertal gilts. Embryo development was observed and the total cell number in each blastocyst generated under the culture conditions was quantified following supplementation of the medium. The embryonic development rate of the blastocyst and hatched blastocyst was higher, but not significantly so, when 0.4% MEM vitamins were supplemented to the in vitro maturation medium of the porcine oocyte. Interestingly, the total number of cells in the blastocyst was significantly higher in the in vitro maturation MEM vitamins supplemented group compared to either the untreated group or the group which had MEM vitamins supplemented to both in vitro maturation and in vitro culture medium simultaneously (p<0.05). Therefore, the supplementation of 0.4% MEM vitamins to the in vitro mature medium has a beneficial effect on the embryonic development of in vitro produced blastocysts derived from the immature porcine oocytes.

In Vitro Growth of Preantral Follicle and Maturation of Intrafollicular Oocyte from Aged Mice

  • Yoon, Jung-Ah;Choi, Jung-Kyu
    • Journal of Animal Reproduction and Biotechnology
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    • v.34 no.1
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    • pp.35-39
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    • 2019
  • This study aimed to recover the ovarian function through in vitro culture of preantral follicles from aged mice. First, we isolated the preantral follicles from ovaries of sixty-seven-week old B6D2F1 mice with decreased fecundity to know how many follicles were present in them, which was 6 preantral follicles including 2 primary, 2 early secondary and late secondary follicles from 8 aged mice. It was confirmed that a few follicles (~2) were present in aged mice through histological analysis compared to adult mice as control. The 9 days of in vitro culture of preantal follicles showed in vitro growth and induced maturation after treatment with hCG (2.5 IU/mL) and EGF (5 ng/mL). Cumulus cells in the cumulus-oocyte complexes (COCs) were removed using hyaluronidase and oocytes at the germinal vesicle (GV) and GV breakdown (GVBD) were obtained from preantral follicle culture of aged mice in vitro. In conclusion, these observations demonstrated that there still were a few preantral follicles in the ovaries of 67 week-old mice, which we were able to culture in vitro and oocytes were obtained from them. This study proposed an in vitro culture system using preantral follicle as a therapeutic strategy for fertility preservation in humans for assisted reproductive medicine.

Effects of Hormones, Oviduct Epithelial Cells, Cumulus Cells during the In Vitro Culture in Medium on In Vitro Developmental Rates of Bisected Bovine Embryos (소 분할배와 호르몬, 난관상피세포, 난구세포와의 공배양이 체외발생율에 미치는 영향에 관한 연구)

  • 김상근;남윤이;이명헌;이만휘
    • Korean Journal of Animal Reproduction
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    • v.21 no.3
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    • pp.281-285
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    • 1997
  • The study was conducted to investigate on in vitro developmental rates of bisected bovine embryos co-culture in 10% FCS+TCM-199 media containing hormones, oviductal epithelial cells and cumulus cells 0 to 7 days after bisection. In vitro developmental rates was defined as development rates on in vitro culture or FDA-test. The results are summarized as follows : 1. In vitro developmental rates of bisected bovine embryos co-cultured in 10% FCS+TCM-199 media containing PMSG+hCG, PMSG+$\beta$-estradiol, hCG+$\beta$-estradiol, PMSG, hCG 0 to 3 days and 4 to 7 days were 16.7~30.0% and 11.1~25.0%, respectively. In vitro developmental rates of bisected embryos co-cultured in 10% FCS+TCM-199 media containing hormones significantly higher than that of non co-culture. 2. In vitro developmental rates of bisected bovine embryos co-cultured 10% FCS+TCM-199 media containing oviductal epithelial cells 0 to 3 days and 4 to 7 days were 25.0% and 22.2%, respectively.

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Effects of Culture Duration, Follicle Stimulating Hormone (FSH) Type, and Activin A Concentration on In Vitro Growth of Preantral Follicles and Maturation of Intrafollicular Oocytes

  • Choi, Jung Kyu
    • Journal of Animal Reproduction and Biotechnology
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    • v.34 no.2
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    • pp.117-122
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    • 2019
  • The objective of this study was to establish an in vitro culture system for ovarian preantral follicles of B6D2F1. First, we optimized the in vitro preantral-follicle culture by culture duration, follicle stimulating hormone (FSH) type, and activin A concentration. Duration of in vitro culture for 9, 11, and 13 days was sufficient for the normal development of preantral follicles to antral follicles. Formation of cumulus cell-oocyte complex (COC) was induced by treatment with human chorionic gonadotropin (hCG; 2.5 IU/mL) and epidermal growth factor (EGF; 5 ng/mL). In addition, metaphase II (MII) oocytes formed during this in vitro culture of preantral follicles. In vitro preantralfollicle culture for 9 days showed higher rates of growth and maturation, thus yielding a greater number of antral follicles, and there were significant differences (p < 0.05) in the number of MII oocytes (that formed from these preantral follicles via differentiation) between the 9-day culture and 11-day or 13-day culture. The follicles cultured for 9 days contained a tightly packed well-defined COC, whereas in follicles cultured for 11 days, the COC was not well defined (spreading was observed in the culture dish); the follicles cultured for 13 days disintegrated and released the oocyte. Second, we compared the growth of the preantral follicles in vitro in the presence of various FSH types. There were no significant differences in the growth and maturation rates and in differentiation into MII oocytes during in vitro culture between preantral follicles supplemented with FSH from Merck and those supplemented with FSH from Sigma. To increase the efficiency of MII oocyte formation, the preantral follicles were cultured at different activin A concentrations (0 to 200 ng/mL). The control follicles, which were not treated with activin A, showed the highest rate of differentiation into antral follicles and into MII oocytes among all the groups (0 to 200 ng/mL). Therefore, activin A (50 to 200 ng/mL) had a negative effect on oocyte maturation. Thus, in this study, we propose an in vitro system of preantral-follicle culture that can serve as a therapeutic strategy for fertility preservation of human oocytes for assisted reproductive medicine, for conservation of endangered species, and for creation of superior breeds.

Study on the In Vitro Maturation and Sperm Penetration Rates of Canine Oocytes

  • Park, Ji-Hoon;Seok, Ho-Bong;Kim, Sang-Keun
    • Reproductive and Developmental Biology
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    • v.34 no.1
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    • pp.21-25
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    • 2010
  • The purpose of this study was to investigate the effects of the collection time, co-culture and sperm penetration of canine oocytes on in vitro maturation and fertilization. The oocytes were cultured in TCM-199 media containing hormonal supplements (10% FCS, 10 IU/ml HCG, 10 IU/ml PMSG) at 5% $CO_2$, 95% air, $38^{\circ}C$. The in vitro maturation rate to MII stage of in vitro oocytes recovered from ovaries that collected at follicular, luteal and inactive phases of the reproductive phase for 44~72 hrs were 19.2%, 12.2%, and 6.0%, respectively. Follicular phases oocytes had a significantly higher in vitro maturation rate than oocytes collected at luteal and anestrus stage (p<0.05). The in vitro maturation rates to the MII stage of canine oocytes after 48 hrs of culture with glutathione, pyruvate, or glutathione + pyruvate were 12.5%, 10.7%, and 17.5%, respectively. This was higher than that in both alone or the combination of the two compared to the control group (19.0%). The sperm penetration rates of in vitro matured oocytes by fresh and frozen semen were 29/80 (36.3%) and 18/80 (22.5%), respectively. Although there are limited reports about canine oocytes co-culture and in vitro fertilization, our results on in vitro maturation is comparable to the results from other researches.

Composition of Culture Medium and Culture Conditions for In vitro Culture of Rice Panicle

  • Lee, Jin-Ho;Lee, Ho-Jin
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.45 no.1
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    • pp.32-37
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    • 2000
  • The in vitro culture of rice panicles is a culturing technique only panicle without other organs in culture solution containing organic substance, so that would be useful to study how assimilate supply affects grain development and maturation. To find the optimum stage for in vitro culture, rice panicles grown in greenhouse were sampled periodically after anthesis and cultured in nutrient medium. The panicles older than 1 weeks after anthesis had produced normal grains. Grain-filling was apparently dependent upon sucrose concentration (8-12 %) in medium, but not affected by nitrogen concentration supplied with glutamine. As far as rice panicle was supplied with sucrose and N in nutrient medium, grains continued accumulation of dry matter and maturation regardless to light condition. Considerably, grain-filling was improved when panicles were positioned horizontally inside flask, so that each grain was partially submerged to nutrient medium.

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In vitro Nasal Cell Culture Systems for Drug Transport Studies

  • Cho, Hyun-Jong;Termsarasab, Ubonvan;Kim, Jung-Sun;Kim, Dae-Duk
    • Journal of Pharmaceutical Investigation
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    • v.40 no.6
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    • pp.321-332
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    • 2010
  • Growing interest in the nasal route as a drug delivery system calls for a reliable in vitro model which is crucial for efficiently evaluating drug transport through the nasal cells. Various in vitro cell culture systems has thus been developed to displace the ex vivo excised nasal tissue and in vivo animal models. Due to species difference, results from animal studies are not sufficient for estimating the drug absorption kinetics in humans. However, the difficulty in obtaining reliable human tissue source limits the use of primary culture of human nasal epithelial cells. This shortage of human nasal tissue has therefore prompted studies on the "passage" culture of nasal epithelial cells. A serially passaged primary human nasal epithelial cell monolayer system developed by the air-liquid interface (ALI) culture is known to promote the differentiation of cilia and mucin gene and maintain high TEER values. Recent studies on the in vitro nasal cell culture systems for drug transport studies are reviewed in this article.