scholarly journals Supplementation of Follicle Stimulating Hormon Into In vitro Maturation Medium to Increase Oocytes Maturation and 4 Cell Stadium Embryo Development of Bligon Goat

2018 ◽  
Vol 42 (2) ◽  
Author(s):  
Yanuar Achadri ◽  
Sigit Bintara ◽  
Diah Tri Widayati
Keyword(s):  
Tissue Culture ◽  
Embryo Development ◽  
Embryo Culture ◽  
In Vitro Maturation ◽  
Culture Medium ◽  
Tissue Culture Medium ◽  
Cleavage Rate ◽  
Maturation Medium ◽  
Phosphate Buffered Saline

The study was carried out to investigate the effect of follicle stimulating hormon (FSH) into in vitro maturation medium to increase oocytes maturation and 4 cell stadium embryo development of Bligon goat. Goat ovaries were obtained from a slaughterhouse and transported to the laboratory in a flask of NaCl at temperature of 31 – 34°C. Oocytes were aspirated from 2 – 6 mm of follicles into a 3 mL syringe (23G needle) that contained Dulbecco’s Phosphate-Buffered Saline. Oocytes were divided into three groups, i.e tissue culture medium (TCM) with FSH supplementation 0, 50, and 100 IU/mL. Oocytes were put into those medium and incubated on 39°C, 5% CO2, and 95% humidity for 24 hours. Matured oocytes were fertilized with capacitated frozen thawed-semen and incubated on 39°C, 5% CO2, and 95% humidity for 5 hours. Fertilized oocytes were washed for 3 times in TCM and incubated in the same condition for embryo culture. The data of FSH supplementation and embryo development were analyzed using randomized completely one way classification. The results showed that the percentages of mature oocytes from FSH supplementation 0, 50, and 100 IU/mL were 70,48±23,22, 78,48±15,80, and 80,29±12,86%, respectively. Cleavage rate of the two cells stage were 36,00±14,22, 44,00±33,94, and 57,45±31,78%, respectively, and for the 4 cells stage were 27,33±22,04, 35,33±40,73, and 39,45±20,38%. It is concluded that supplementation of FSH in the maturation medium could not increase the percentages of in vitro maturation and embryo development.

150 DEVELOPMENT OF ZONA-FREE AND WITH ZONA PARTHENOGENETIC GOAT EMBRYOS IN DIFFERENT MEDIA

2010 ◽  
Vol 22 (1) ◽  
pp. 233
Author(s):  
M. K. Jena ◽  
D. Malakar ◽  
A. K. De ◽  
S. Garg ◽  
Y. S. Akshey
Keyword(s):  
Embryo Development ◽  
Chemical Activation ◽  
Formation Rate ◽  
Electrical Activation ◽  
Cleavage Rate ◽  
Maturation Medium ◽  
Parthenogenetic Embryo ◽  
Oviductal Fluid ◽  
Phosphate Buffered Saline

The present study was carried out to see the developmental efficiency of zona-free and with zona parthenogenetic goat embryos cultured in Research Vitro Cleave from Cook Australia (RVCL), Embryo Development Media (EDM), modified synthetic oviductal fluid (mSOF), and modified Charles Rosenkrans media (mCR2a). Zona-free embryos were cultured in 4 media, whereas with zona embryos were cultured in 3 media except mCR2a. Ovaries were collected from slaughterhouse and oocytes were isolated by puncturing the follicles in medium containing Dulbecco’s phosphate-buffered saline, 3% BSA, and 50 μg mL-1 gentamicin. Oocytes were matured in maturation medium containing TCM-199 (HEPES modified), 0.05 mg mL-1 Na pyruvate, 0.003 mg mL-1 L-glutamine, 5.5 mg mL-1 glucose, 3 mg mL-1 BSA, 5 μg mL-1 FSH, 10 μg mL-1 LH, 1 μg mL-1 estradiol-17β, 50 μg mL-1 gentamicin, and 10% FBS in 5% CO2 in air at 38.5°C. The COC (15 to 20 oocytes) were placed in 100-μL droplets of maturation medium and incubated in a CO2 incubator (5% CO2 in air) with maximum humidity at 38.5°C for 27 h. Matured oocytes were made cumulus free by treatment with hyaluronidase (0.5 mg mL-1) and zona-free by pronase (2 mg mL-1) in zona-free parthenogenesis. Then the oocytes were activated by 5 μM Ca ionophore for 5 min in a CO2 incubator and then treated with 2 mM 6-DMAP for 4 h. Activation was also done by electrical activation with DC 1.78 kV cm-1, 20 μs, and 2 pulses. Then the zona-free oocytes were kept for in vitro culture in 4 types of media such as RVCL, EDM, mSOF, andm CR2a for 7 days in 5% CO2 in air at 38.5°C. The cleavage rate andmorulae formation were observed in RVCL 40.95%, 13.95%, in EDM 46.92%, 14.75%, in mCR2a 56.66%, 5.88%, and in mSOF 48.23%, 14.63%, respectively. The cleavage rate and morulae formation were also found 55.9%, 14.63% during chemical activation and 32%, 12.5% in electrical activation. Hence, better result was found in chemical activation than electrical activation. For with zona parthenogenesis, the matured oocytes were chemically activated by 5 μM Ca ionophore for 5 min and 2 mM 6-DMAP for 4 h. Then the oocytes were cultured in RVCL, EDM, and mSOF in 100-μL micro-drops media for 7 days. The cleavage, morulae, and early blastocyst production rate were as follows: cleavage rate 75.68%, 72.03%, and 57.11%; morulae 44.61%, 30.29%, and 40.22%; and early blastocyst 17.49%, 11.88%, and 25.01% in RVCL, EDM, and mSOF, respectively. Hatched blastocyst formation rate was 6.75%, 5.48%, and 1.15% in RVCL, EDM, and mSOF, respectively. It could be concluded that zona-free parthenogenetic embryos were produced better in EDM medium and with chemical activation. With zona parthenogenetic embryo development was significantly (P < 0.05) higher in RVCL and EDM media.

scholarly journals PENGGUNAAN CAIRAN FOLIKEL DALAM MEDIA MATURASI IN VITRO OOSIT KAMBING BLIGON

2014 ◽  
Vol 8 (1) ◽  
Author(s):  
Diah Tri Widayati ◽  
Devita Hery Fatmawati ◽  
Nony Ariesta ◽  
Kustono K
Keyword(s):  
Tissue Culture ◽  
Culture Medium ◽  
Tissue Culture Medium ◽  
Phosphate Buffered Saline

Penelitian ini bertujuan mengetahui pengaruh medium maturasi terhadap kompetensi perkembangan in vitro oosit kambing bligon. Ovarium kambing diperoleh dari Rumah Potong Hewan, dibawa ke laboratorium dalam 0,9% NaCl pada suhu 31-34 C. Oosit diaspirasi dari folikel berdiameter 2-6 mm dengan menggunakan syringe 3 ml dan jarum 23 G. Oosit dengan minimal 2 lapisan sel-sel kumulus diproses untuk maturasi in vitro. Oosit dicuci 3 kali dengan dulbecco’s phosphate buffered saline (DPBS) dan tissue culture medium 199 (TCM) 199, kemudian didistribusikan ke dalam 50 μl medium maturasi yang berbeda, yaitu TCM 199 (tanpa suplementasi),

148 Effect of concentration of methionine in maturation and culture medium on cleavage rate of oocytes from alpaca (Lama pacos)

2019 ◽  
Vol 31 (1) ◽  
pp. 199
Author(s):  
M. L. Uchuari ◽  
M. Artica ◽  
J. C. Villanueva ◽  
W. F. Huanca ◽  
W. Huanca
Keyword(s):  
In Vitro Maturation ◽  
Culture Medium ◽  
Egg Yolk ◽  
Cumulus Cells ◽  
Cleavage Rate ◽  
Sodium Pyruvate ◽  
Maturation Medium ◽  
Thermos Flask ◽  
Lama Pacos

Maturation time of oocytes from alpacas is around 38 to 40h (Huanca et al. 2009) that would induce an increase in reactive oxygen species during in vitro maturation and IVF and cause cytotoxic damage to gametes. The objective of this study was to determine the optimal concentration of methionine during in vitro maturation on cleavage rate of alpacas oocytes following IVF. Cumulus-oocyte complexes were collected from slaughterhouse ovaries and transported in a thermos flask containing a saline solution 0.9% and antibiotic, antimycotic at 35°C. Cumulus-oocyte complexes were aspirated from follicles &gt;2mm and evaluated with a stereomicroscope for selection. Only cumulus-oocyte complexes with a homogeneous cytoplasm and with 2 or more layers of cumulus cells were selected to be cultured in maturation medium TCM-199 supplemented with 10% FCS (v:v) plus 0.5μg mL−1 FSH, 10μg mL−1 hCG, 0.2mM sodium pyruvate, 50μg mL−1gentamycin and 1μg mL−1 oestradiol under mineral oil by 38h. Testes of mature males were collected from a slaughterhouse and transported to the laboratory. Caudal epididymide was isolated, and fluid, rich in spermatozoa, was aspirated in syringes containing 2mL of Tris-fructose-egg yolk extender. Motile spermatozoa were obtained by centrifugation at 700×g in a Percoll discontinuous gradient (22.5: 45.0%) for 10min. The supernatant was removed by aspiration, and the pellet was resuspended in TL stock and centrifuged again at 700×g for 5min. Spermatozoa and oocytes were co-incubated by 18h at 39°C with 5% CO2. Presumptive zygotes were culture in KSOMaa medium and evaluated at 72h. The treatments include 0, 14 and 21 μM of methionine in maturation and culture medium. Data were analysed by ANOVA, and results are presented in Table 1. The results suggest that addition of methionine in maturation and culture medium improve the cleavage rate in oocytes from alpacas. Table 1.Cleavage rate (%) following in vitro maturation at different concentrations of methionine Proyect 405-PNICP-PIAP-2014, INNOVATE-PERU, is acknowledged.

Cysteamine supplementation of in vitro maturation medium, in vitro culture medium or both media promotes in vitro development of buffalo (Bubalus bubalis) embryos

2008 ◽  
Vol 20 (2) ◽  
pp. 253 ◽  
Author(s):  
T. Anand ◽  
D. Kumar ◽  
M. S. Chauhan ◽  
R. S. Manik ◽  
P. Palta
Keyword(s):  
In Vitro Culture ◽  
In Vitro Maturation ◽  
Culture Medium ◽  
Bubalus Bubalis ◽  
Cell Number ◽  
Hatching Rate ◽  
Cleavage Rate ◽  
Maturation Medium ◽  
Vitro Development

The effects of supplementation of in vitro maturation (IVM) or in vitro culture (IVC) or both IVM and IVC media with cysteamine on the yield, hatching rate (HR) and total cell number (TCN) of buffalo blastocysts were examined. Oocytes obtained from slaughterhouse buffalo ovaries were subjected to IVM and IVF. The IVM or IVC media were supplemented with 0, 50, 100 or 200 µm cysteamine. Supplementation of IVM medium with 50 µm cysteamine increased (P < 0.01) the cleavage rate and blastocyst yield without affecting the HR and TCN whereas a higher concentration of 200 µm significantly (P < 0.05) reduced the blastocyst yield but not TCN. Similar increases in blastocyst yield, without any effect on HR and TCN were observed after supplementation of the IVC medium with 100 (P < 0.01) or 50 µm (P < 0.05) cysteamine, whereas 200 µm cysteamine was ineffective. Supplementation of both IVM medium with 50 µm cysteamine and of IVC medium with 100 µm cysteamine increased the yield of blastocysts and hatched blastocyst by over 100% (P < 0.01) compared with the controls without any adverse effects on HR or TCN. The results of the present study suggest that supplementation of both IVM and IVC media improves the yield of blastocysts without compromising their health.

scholarly journals Comparison of Quinn's Advantage fertilization medium and tissue culture medium 199 for in vitro maturation of oocytes

2014 ◽  
Vol 53 (1) ◽  
pp. 17-20 ◽  
Author(s):  
Yu-Hung Lin ◽  
Jiann-Loung Hwang ◽  
Lee-Wen Huang ◽  
Kok-Min Seow ◽  
Bih-Chwen Hsieh ◽  
...  
Keyword(s):  
Tissue Culture ◽  
In Vitro Maturation ◽  
Culture Medium ◽  
Tissue Culture Medium

scholarly journals Suplementasi Hormon Gonadotropin Pada Medium Maturasi In Vitro Untuk Meningkatkan Perkembangan Embrio Stadium 4 Sel Kambing Bligon

2016 ◽  
Vol 40 (2) ◽  
pp. 83 ◽  
Author(s):  
Dewi Pranatasari ◽  
Kustono Kustono ◽  
Diah Tri Widayati
Keyword(s):  
Embryo Development ◽  
In Vitro Maturation ◽  
Spherical Shape ◽  
Maturation Stage ◽  
Embryo Morphology ◽  
Cleavage Rate ◽  
Vitro Fertilization ◽  
Maturation Medium ◽  
Cell Embryo

The study was carried out to investigate the effect of gonadotropin hormone supplementation into in vitro maturation medium on maturation, fertilization and embryo development of Bligon goats. This research steps consist of oocyte collection, in vitro maturation, in vitro fertilization, and in vitro embryo development. At the maturation stage the oocyte that had been collected and divided into two groups based on the maturation medium, that was tissue culture medium (TCM) with supplementation of GnRH 0 IU/mL and GnRH 25 IU/mL. Oocyte and embryo morphology data were analyzed descriptively. Maturation rate and embryo development data were analyzed by using independent sample t-test. Fertilization data was analyzed descriptively. The result showed the percentages of mature oocytes from gonadotropin supplementation of 0 IU/mL and 25 IU/mL were 54.10±25.97 and 54.89±26.44%, respectively. Expansion cumulus cells surrounding the oocytes might indicated the mature oocytes. Cleavage rate of the 2 cells stage were 13,02±11,09 and 27,01±16,65%; respectively, and for the 4 cells stage were 10,16±10,01% and 16,67±14.91%. Embryos obtained from the treatment, indicated uniform of blastomeres in the size, tight, compact, intact, and round-spherical shape. It could be concluded that supplementation of gonadotropin hormone into in vitro maturation medium could not increase the rate of oocyte maturation and 4 cell embryo development, but it could increase 2 cell embryo development of Bligon goats. Hormone supplementation could improved the maturation and embryo quality.

Supplemented tissue culture medium 199 is a better medium for in vitro maturation of oocytes from women with polycystic ovary syndrome women than human tubal fluid

2009 ◽  
Vol 91 (2) ◽  
pp. 509-513 ◽  
Author(s):  
Carlos Henrique Medeiros de Araujo ◽  
Daniela Nogueira ◽  
Maria Cristina Picinato Medeiros de Araujo ◽  
Wellington de Paula Martins ◽  
Rui Alberto Ferriani ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Polycystic Ovary Syndrome ◽  
In Vitro Maturation ◽  
Culture Medium ◽  
Polycystic Ovary ◽  
Tissue Culture Medium ◽  
Ovary Syndrome ◽  
Human Tubal Fluid

scholarly journals Effect of adding growth factors during in vitro maturation on the developmental potentials of ewe oocytes selected by brilliant cresyl blue staining

2021 ◽  
Vol 14 (2) ◽  
pp. 452-456
Author(s):  
Mohamed Fathi ◽  
Amr F. Elkarmoty
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Embryo Development ◽  
In Vitro Maturation ◽  
Blue Staining ◽  
Brilliant Cresyl Blue ◽  
Cresyl Blue ◽  
Nuclear Maturation ◽  
Maturation Medium

Aim: Several factors had been concerned with the developmental competence of the sheep oocyte. This study aims to investigate the effect of adding growth factors (insulin-like growth factor 1 [IGF-1] and epidermal growth factor [EGF]) in the maturation medium of ewe oocytes selected based on brilliant cresyl blue (BCB) screening on in vitro maturation (IVM), fertilization, and pre-implantation embryo development. Materials and Methods: Cumulus-oocyte complexes (COCs) were obtained from the ovaries of slaughtered ewes by either aspiration or slicing techniques. COCs were in vitro matured in a medium containing IGF-1 and EGF (control group). For BCB screening, oocytes were stained and divided into BCB+ oocytes that matured in the same maturation conditions without adding growth factors (Group 2) or in the presence of growth factors (Group 3), and BCB– oocytes that matured in medium without growth factors (Group 4) or with growth factors (Group 5). Results: The supplementation of the maturation medium with growth factors during IVM of (BCB+) oocytes resulted in a significant increase in nuclear maturation rate (90.9%), fertilization rate (75.6%), and embryo developmental rates (60.0%, 46.7%, and 33.3% for cleavage, morula, and blastocyst, respectively). Conclusion: Culturing BCB+ oocytes in a maturation medium containing both EGF and IGF-1 showed a significant improvement in nuclear maturation, fertilization, and pre-implantation embryo development in vitro.

Effects of changing culture medium on preimplantation embryo development in rabbit

Zygote ◽  
2021 ◽  
pp. 1-6
Author(s):  
Haixia Wang ◽  
Wenbin Cao ◽  
Huizhong Hu ◽  
Chenglong Zhou ◽  
Ziyi Wang ◽  
...  
Keyword(s):  
Embryo Development ◽  
Embryo Culture ◽  
Culture Medium ◽  
Cell Number ◽  
Apoptotic Index ◽  
Total Cell ◽  
Toxic Substances ◽  
Total Cell Number ◽  
Preimplantation Embryo Development

Summary Many studies have focused on the optimization of the composition of embryo culture medium; however, there are few studies involving the effect of a culture medium changing procedure on the preimplantation development of embryos. In this study, three groups were designed: a non-renewal group, a renewal group and a half-renewal group. The levels of reactive oxygen species (ROS), apoptotic index, blastocyst ratio and blastocyst total cell number were analyzed in each group. The results showed that the ROS level and the apoptotic index of blastocyst in the non-renewal group were significantly higher than in the renewal group and the half-renewal group (P < 0.05). The blastocyst ratio and blastocyst total cell number were significantly higher in the half-renewal group than that in non-renewal group and the renewal group (P < 0.05). These results demonstrated that the procedure of changing the culture medium influenced ROS level, apoptotic index, blastocyst ratio and total cell number of blastocysts. In addition, the result suggested that changing the culture medium may lead to a loss of important regulatory factors for embryos, while not changing the culture medium may lead to the accumulation of toxic substances. Half-renewal can alleviate the defects of both no renewal and renewal, and benefit embryo development. This study will be of high value as a reference for the optimization of embryo culture in vitro, and is very significant for assisted reproduction.

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