• Journal of Embryo Transfer
• /
• v.17 no.2
• /
• pp.145-151
• /
• 2002

The aim of these experiments was to investigate in vitro nuclear maturation of canine oocyte collected from various stages of estrus cycles, Ovaries were obtained from 1 to 4 year-old mongrel bitch and minced for oocyte collection in phosphate buffered saline with 100 iu penicillin-G $m\ell$／sup -1／, 50 $\mu\textrm{g}$ streptomycin sulphate $m\ell$／sup -1／ and 0.1％ (w／v) polyvinyl alcohol. Cumulus-oocyte-complexes (COCs) were washed in HEPES buffered tissue culture medium (TCM)199 and in vitro matured in TCM-199 culture medium supplemented with sodium pyruvate 0.028mg/$m\ell$, L-glutamine 0.146mg／$m\ell$, penicillin G 10,000 IU／$m\ell$, streptomycin 0.031mg／$m\ell$ and 10％ (v／v) fatal calf serum. COCs were in vitro matured for 48～72 hrs at 39$^{\circ}C$ in humidified 5％ $CO_2$ in air atmosphere. In vitro matured oocytes were remove the cumulus cells using 0.2％ (v／v) hyaluronidase. After denuding, oocyte were placed in acetic acid : methanol : chlorform solusion (3:6 : 1.5 v／v) for 30 sec and acetic acid: ethanol(1:3 v／v) for 48hrs fixation. Nuclear maturation was classified to GV, GVBD, MI, MII and degenerate oocyte under microscopy after 1％ aceto-orcein stain. In vitro maturation rates at 48hrs were not significantly difference among the oocytes collected from different stage of estrus at 15.9％, 16.3％, 23.7％ and 18.2％ for anestrus, proestrus, estrus and diestrus. However, the oocytes maturation(36.6％) of collected from estrus ovaries were significantly different from oocytes derived from proesturs, diestrus and anestrus ovaries(30.8％, 17.5％ and 22.1％; p＜0.05). The overall in vitro maturation rates were significantly higher (p＜0.05) in 72hrs culture than 48hrs culture system. In summary, there was a tendency for higher in vitro maturation rates with the oocyte collected from estrus ovary than other stages of estrus. Also, for nuclear maturation, in vitro culture of oocyte for 72hrs was better than 48hrs culture.

• Clean Technology
• /
• v.12 no.3
• /
• pp.165-170
• /
• 2006

When ammonia ($NH_3$) and hydrogen sulfide ($H_2S$) in binary mixture gases were supplied to a biofilter packed with biomedia made of polyurethane, PVA, and worm cast. No odor gases were detected on the outlet of the biofilter when $NH_3$ and $H_2S$ were separately supplied to the biofilter at space velocity(SV) of $50h^{-1}$ until inlet $NH_3$ concentration was increased up to 300 ppmv and inlet $H_2S$ to 428 ppmv. While, inlet $NH_3$ concentration maintained at 50 ppmv, inlet $H_2S$ concentration increased from 1 to 489 ppmv, and the removal efficiency of each gas was investigated. After that, $NH_3$ concentration increased step by step such as 80, 100, 200, 300, 400 and 500 ppmv. $H_2S$ concentration increased gradually when $NH_3$ concentration was set up at each condition. Under each condition, removal efficiency of $NH_3$ and $H_2S$ gas was investigated by analysing the gases sampled from the inlet and outlet of the biofilter. When binary gases were supplied to the biofilter and inlet $NH_3$ concentration was increased from 50 to 300 ppmv, elimination capacity of $NH_3$ increase linearly as inlet loading increased to $11.14g\;N{\cdot}m^{-3}{\cdot}h^{-1}$. However, as inlet $NH_3$ concentration increased over 300 ppmv, both removal efficiency and elimination capacity decreased while inlet loading increased. $H_2S$ removal efficiency was not affected seriously by the simultaneous supply of $NH_3$ when maximum inlet loading of $H_2S$ was under $40.27S{\cdot}m^{-3}{\cdot}h^{-1}$ and maximum inlet loading of $NH_3$ was under $15.25N{\cdot}m^{-3}{\cdot}h^{-1}$.

• Journal of Embryo Transfer
• /
• v.21 no.2
• /
• pp.157-162
• /
• 2006

It is well known that unidentified factors in sera, hormones and growth factors promote the proliferation of granulosa cells and nuclear maturation of bovine COCs (cumulus oocytes complexes) in vitro. Attempts had been developed the simple composition of culture media and similar system to in vivo conditions has been applied. In the present study, we investigated the effect of FGF (fibroblast growth factor) on in vitro maturation and in vitro development of Hanwoo COCs. When the COCs were matured in HPM 199 (Inst. of Functional peptide, Japan) containing 0.1, 1 and 10 ng/ml FGF for 24 hr, maturation rates to metaphase II ($70.0{\sim}75.0%$) were significantly higher (p<0.05) than that of control group (0 ng/ml FGF, 37.5%). When matured COCs with FGF were cultured in maturation medium after in vitro fertilization, developmental rates to blastocysts were 9.5, 0 and 2.9%, respectively, compared to 25.0% of the control group (p<0.05). When the matured COCs with FGF were cultured in HPM 199 (IFP971, Inst. of Functional peptide, Japan) containing 10% FBS, 0.8% BSA or 0.1% PVA (polyvinyl alcohol), the blastocyst formation rates were 12.4, 12.8 and 8.5%, respectively, while the rates of matured COCs with FGF and cultured with IVMD and IVD (Inst. of Functional peptide, Japan) without serum were 38.4% and 34.8%, respectively (p<0.05). These results suggested that FGF is available for in vitro maturation of bovine COCs and is not suitable for in vitro development, but further investigation would be need for finding the synergistic autocrine/paracrine fashion of other growth factors in early bovine embryo development.