scholarly journals 87 VITRIFICATION OF BOVINE OOCYTES TREATED WITH CHOLESTEROL-LOADED METHYL-β-CYCLODEXTRIN

2005 ◽  
Vol 17 (2) ◽  
pp. 193
Author(s):  
G. Horvath ◽  
L. Solti ◽  
G. Seidel
Keyword(s):  
Fatty Acid ◽  
Defined Medium ◽  
Chemically Defined Medium ◽  
Chemically Defined Media ◽  
Defined Media ◽  
Standard Procedures ◽  
Maturation Medium ◽  
Cell Embryo ◽  
Bovine Oocytes ◽  
Better Than

Addition of cholesterol-loaded cyclodextrin (CLC) can increase sperm cryosurvival (Purdy et al. 2000 Cryobiology 48, 36–45). The purpose of this study was to determine if cryosurvival of vitrified oocytes could be improved by incubation with CLC prior to vitrification. Slaughterhouse-derived cumulus oocyte complexes were matured in a chemically defined medium with fatty acid-free BSA and hormones for 21 h followed by partial cumulus removal with 100 U/mL hyaluronidase and gentle pipetting. For an additional hour, oocytes were placed into maturation medium supplemented with 0.5% PVA instead of BSA with or without 2.5 mg/mL CLC. At 22 h after the start of maturation, oocytes were transferred to handling media containing 20% FCS or 0.5% PVA in TCM-199 + HEPES (HTCM-199). Oocytes with approximately 3 layers of cumulus were vitrified in two steps. First, they were exposed to VS1 (10% ethylene glycol (EG), 10% DMSO, 6% PVP, or 20% FCS, in HTCM-199) for 30 s, then exposed to VS2 (20% EG, 20% DMSO, 6% PVP, or 20% FCS, 0.48 M galactose in HTCM-199) for 25 s, loaded into cryoloops in groups of five, and plunged into liquid nitrogen. Rapidly warmed oocytes were moved stepwise through 0.5, 0.25, 0.125, and 0 M galactose in HTCM-199 + 20% FCS, 3 min each. All procedures were conducted at 39°C. Warmed oocytes were placed in maturation medium for an additional hour, fertilized with semen from 3 bulls, 3 replicates each, and cultured according to standard procedures (Zhang et al. 2003 Theriogenology 60, 1657–1663). For each replicate, 30 oocytes were assigned to the following treatments: A: chemically-defined media with PVA for the last hour of maturation, handling and vitrification; B: same as A except CLC treatment, for 1 h before vitrification; C: chemically defined media for maturation, but with 20% FCS for HM, VS1 and VS2. Data were analyzed by ANOVA. CLC treatment resulted in higher cleavage rates and 8- to 16-cell embryo production, but not higher blastocyst (bl) production (Table 1). Non-vitrified oocytes developed better than vitrified ones (means: cleavage, 76%; 8- to 16-cell, 64%; bl D8, 21%; bl D9, 24%). Further studies with vitrification of cholesterol-loaded cyclodextrin-treated oocytes and chemically defined media are warranted. Table 1. Development of vitrified oocytes (LS means ± SE)

Cultivation and properties of Neisseria sp. grown in chemically defined media

1972 ◽  
Vol 18 (7) ◽  
pp. 1087-1090 ◽  
Author(s):  
C. P. Kenny ◽  
B. B. Diena ◽  
R. Wallace ◽  
L. Greenberg
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Present Report ◽  
Defined Medium ◽  
Chemically Defined Medium ◽  
Chemically Defined Media ◽  
Defined Media ◽  
Specific Antisera

Neisseria Chemically Defined Medium (NCDM) has been used routinely in our laboratory for a variety of purposes. The present report describes the development of NCDM agar, wherein the NCDM base is sterilized by filtration and defined supplements and agar are added. The medium is transparent and both meningococci and gonococci grow within 72 h. When grown on NCDM agar, Types 2 and 3 gonococcal colonies tend to revert to Type 1. The serological grouping of meningococci with specific antisera is not affected by growth on this medium.Parallel investigations on the growth of these species in liquid NCDM demonstrated that the yield of Neisseria gonorrhoeae is enhanced when the medium is sterilized by filtration.

scholarly journals Effect of methionine and cysteine deprivation on growth of different natural isolates of Lactobacillus spp. in chemically defined media

2008 ◽  
Vol 60 (4) ◽  
pp. 509-517 ◽  
Author(s):  
Jelena Lozo ◽  
Jelena Begovic ◽  
B. Jovcic ◽  
Natasa Golic ◽  
L. Topisirovic
Keyword(s):  
Amino Acids ◽  
Defined Medium ◽  
Ecological Niches ◽  
Chemically Defined Medium ◽  
Chemically Defined Media ◽  
Defined Media ◽  
Natural Isolates ◽  
Lactobacillus Spp

The purpose of this study was to determine the ability of natural isolates of lactobacilli from different ecological niches to grow in a chemically defined medium in the presence or absence of sulphur-containing amino acids, methionine and/or cysteine. The obtained results indicate that cysteine is essential for growth of L. paracasei subsp. paracasei BGHN14 and BGSJ2-8, while methionine is essential for isolates BGHN40, BGCG31, and BGHV54T of the species L. plantarum. Methionine is also essential for growth of L. rhamnosus BGHV58T. Other analyzed strains, such as L. plantarum BGSJ3-18, BGZB19, BGHV52Ta, and BGHV43T, require the presence of both amino acids for their growth.

scholarly journals 99MACROMOLECULES FOR VITRIFYING BOVINE OOCYTES

2004 ◽  
Vol 16 (2) ◽  
pp. 171
Author(s):  
G. Horvath ◽  
G.E. Seidel
Keyword(s):  
Fetal Calf Serum ◽  
Calf Serum ◽  
Cumulus Cells ◽  
Defined Medium ◽  
Standard Procedures ◽  
Zona Hardening ◽  
Maturation Medium ◽  
Experimental Treatments ◽  
Bovine Oocytes

Vitrification of oocytes would make them available for research and clinical purposes wherever and whenever needed. However, development rates and quality of blastocysts arising from vitrified oocytes have been low. Zona hardening following exposure to vitrification solutions and cooling could contribute to low fertilization rates. Addition of fetal calf serum (FCS) to handling media and vitrification solutions can prevent zona hardening, but FCS may be detrimental to resulting embryos and spread viral diseases, and its composition varies among batches. Fetuin is the component responsible for the protective effect of FCS (Landim-Alvarenga FC et al., 2002 Anim. Reprod. Sci. 71, 181–191). Our objective was to determine whether fetuin is a suitable substitute for FCS during vitrification. Oocytes derived from slaughterhouse ovaries were matured in a chemically defined medium with hormones and with or without 1mgmL−1 fetuin at 39°C in 5% CO2 in air. At 22h after the start of maturation, oocytes were transferred to one of the following handling media: 2% BSA, 2% BSA+1mgmL−1 fetuin, or 20% FCS in TCM-199+HEPES (HTCM-199). In the same media plus 100IUmL−1 hyaluronidase, cumulus cells were partly removed by gentle pipetting. Oocytes with approximately 3 layers of cumulus were chosen for two-step vitrification. First, they were exposed to VS1 (10% ethylene glycol (EG), 10% DMSO, 6% PVP in HTCM-199) for 30s, then to VS2 (20% EG, 20% DMSO, 6% PVP, 0.48M galactose in HTCM-199) for 25s, loaded into cryoloops in groups of five, and plunged into liquid nitrogen. Rapidly warmed oocytes were moved stepwise in 0.5, 0.25, 0.125, and 0M galactose in HTCM-199+20% FCS, 3 min each. All procedures were conducted at 39°C. Warmed oocytes were placed in maturation medium for an additional hour, and then fertilized and cultured according to standard procedures (Olson SE and Seidel GE Jr 2000 J. Anim. Sci. 78, 152–157). About 2000 oocytes were used in 11 replicates with semen of 4 bulls. The experimental treatments and controls were: A: maturation BSA, handling BSA; B: maturation BSA, handling FCS; C: maturation BSA, handling BSA+fetuin; D: maturation BSA+fetuin, handling BSA+fetuin; E: non-vitrified control, maturation BSA; F: non-vitrified control, maturation BSA+fetuin (Table 1). Controls did not differ (P>0.1) from each other, nor were there differences among vitrification treatments. Controls resulted in greater cleavage and more 8-cell embryos and blastocysts than vitrified treatments, and also tended to have higher cell numbers. In summary, fetuin can replace FCS during handling without decreasing the success of vitrification. Table 1 Development of vitrified oocytes and controls

Optimum concentrations of essential components for the development of Angiostrongylus costaricensis in vitro

1996 ◽  
Vol 70 (2) ◽  
pp. 173-175 ◽  
Author(s):  
H. Hata
Keyword(s):  
Young Adult ◽  
Choline Chloride ◽  
Defined Medium ◽  
The Other ◽  
Chemically Defined Medium ◽  
Chemically Defined Media ◽  
Defined Media ◽  
Essential Components ◽  
Third Stage

AbstractThird-stage larvae of Angiostrongylus costaricensis were cultured to young adult stages in Waymouth's chemically defined medium MB 752/l, which comprises higher concentrations of the essential components histidine, lysine, methionine, tryptophan, choline chloride and glucose than various other chemically defined media. The present study has shown that choline chloride and tryptophan are required at relatively higher concentrations for worm development than those of the other essential components.

scholarly journals Unique Host Iron Utilization Mechanisms of Helicobacter pylori Revealed with Iron-Deficient Chemically Defined Media

2010 ◽  
Vol 78 (5) ◽  
pp. 1841-1849 ◽  
Author(s):  
Olga Senkovich ◽  
Shantelle Ceaser ◽  
David J. McGee ◽  
Traci L. Testerman
Keyword(s):  
Helicobacter Pylori ◽  
Defined Medium ◽  
Normal Serum ◽  
Chemically Defined Medium ◽  
Limited Growth ◽  
Defined Media ◽  
Iron Deficient ◽  
Novel Strategy ◽  
H Pylori

ABSTRACT Helicobacter pylori chronically infects the gastric mucosa, where it can be found free in mucus, attached to cells, and intracellularly. H. pylori requires iron for growth, but the sources of iron used in vivo are unclear. In previous studies, the inability to culture H. pylori without serum made it difficult to determine which host iron sources might be used by H. pylori. Using iron-deficient, chemically defined medium, we determined that H. pylori can bind and extract iron from hemoglobin, transferrin, and lactoferrin. H. pylori can use both bovine and human versions of both lactoferrin and transferrin, contrary to previous reports. Unlike other pathogens, H. pylori preferentially binds the iron-free forms of transferrin and lactoferrin, which limits its ability to extract iron from normal serum, which is not iron saturated. This novel strategy may have evolved to permit limited growth in host tissue during persistent colonization while excessive injury or iron depletion is prevented.

scholarly journals In vitro production of cattle blastocysts in chemically defined medium with or without insulin supplementation

1996 ◽  
Vol 5 (5) ◽  
pp. 509-514 ◽  
Author(s):  
Kristiina Bredbacka ◽  
Peter Bredbacka
Keyword(s):  
Bovine Serum Albumin ◽  
Culture Medium ◽  
Defined Medium ◽  
Blastocyst Stage ◽  
Chemically Defined Medium ◽  
In Vitro Production ◽  
Cell Numbers ◽  
Vitro Production ◽  
Bovine Oocytes

In this study we evaluated the use of a chemically defined medium in the production of blastocysts from bovine oocytes fertilized in vitro. As culture medium we used CRI-PVP, a modification of CRlaa medium with bovine serum albumin replaced by polyvinylpyrrolidone. After 168 h of culture (192 h after insemination) 8.7%, 10.5 and 12.8% of the cleaved embryos developed to the blastocyst stage in the presence of 0, 2 or 200 nM insulin, respectively. The supplementation of 200 nM insulin tended to increase cell numbers in morulae and blastocysts (P=0.10). It is concluded that CRI-PVP can be used as a chemically defined medium in the production of blastocysts from bovine 1-cell embryos. However, further modifications are needed, and the insulin concentrations used may be below the optimum for blastocyst production.

scholarly journals Development of Chemically Defined Media to Express Trp-Analog-Labeled Proteins in a Lactococcus lactis Trp Auxotroph

2016 ◽  
Vol 26 (4) ◽  
pp. 269-276
Author(s):  
Jinfeng Shao ◽  
Marcelo F.M. Marcondes ◽  
Vitor Oliveira ◽  
Jaap Broos
Keyword(s):  
Amino Acids ◽  
New Media ◽  
Lactococcus Lactis ◽  
Unnatural Amino Acids ◽  
Defined Medium ◽  
Trna Synthetase ◽  
Growth Media ◽  
Heterologous Proteins ◽  
Chemically Defined Media ◽  
Defined Media

Chemically defined media for growth of <i>Lactococcus lactis</i> strains contain about 50 components, making them laborious and expensive growth media. However, they are crucial for metabolism studies as well as for expression of heterologous proteins labeled with unnatural amino acids. In particular, the <i>L. lactis</i> Trp auxotroph PA1002, overexpressing the tryptophanyl tRNA synthetase enzyme of <i>L. lactis</i>, is very suitable for the biosynthetic incorporation of Trp analogs in proteins because of its most relaxed substrate specificity reported towards Trp analogs. Here we present two much simpler defined media for <i>L. lactis</i>, which consist of only 24 or 31 components, respectively, and with which the <i>L. lactis</i> Trp auxotroph shows similar growth characteristics as with a 50-component chemically defined medium. Importantly, the expression levels of two recombinant proteins used for evaluation were up to 2-3 times higher in these new media than in the 50-component medium, without affecting the Trp analog incorporation efficiency. Taken together, the simplest chemically defined media reported so far for <i>L. lactis</i> are presented. Since<i> L. lactis</i> also shows auxotrophy for Arg, His, Ile, Leu Val, and Met, our simplified media may also be useful for the biosynthetic incorporation of analogs of these five amino acids.

Development of bovine oocytes matured, fertilized and cultured in a serum-free, chemically defined medium

1991 ◽  
Vol 35 (6) ◽  
pp. 1197-1207 ◽  
Author(s):  
Y. Takagi ◽  
K. Mori ◽  
M. Tomizawa ◽  
T. Takahashi ◽  
S. Sugawara ◽  
...  
Keyword(s):  
Defined Medium ◽  
Chemically Defined Medium ◽  
Serum Free ◽  
Bovine Oocytes

The effects of 17β-estradiol and protein supplement on the response to purified and recombinant follicle stimulating hormone in bovine oocytes

Zygote ◽  
2002 ◽  
Vol 10 (1) ◽  
pp. 65-71 ◽  
Author(s):  
Atef Ali ◽  
Marc-André Sirard
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Follicle Stimulating Hormone ◽  
Defined Medium ◽  
Protein Supplement ◽  
Maturation Medium ◽  
17Β Estradiol ◽  
Epidermal Growth ◽  
Bovine Oocytes

During in vitro maturation of bovine oocytes, effects of gonadotropins (FSH, LH) and growth factors such as epidermal growth factor (EGF) vary among studies. Now that we can use defined or semi-defined medium, it becomes possible to evaluate recombinant products to assess their roles. Therefore, this study was designed to evaluate the effect of purified porcine (pFSH) or recombinant human (r-hFSH; 5, 50, or 500 ng/ml) follicle stimulating hormone, luteinising hormone (LH; 50, 500 or 5000 ng/ml) and epidermal growth factor (EGF; 5, 10, 30 or 50 ng/ml) on subsequent embryonic development of in vitro matured bovine oocytes. In addition, the presence of bovine serum albumin (BSA; 8 mg/ml) as a protein supplement during in vitro maturation (IVM) was studied. For all treatments, cumulus-oocyte complexes were matured in defined maturation medium consisting of synthetic oviduct fluid. Addition of LH to the maturation medium at all concentrations studied did not increase the proportion of oocytes developing to the morula and blastocyst stages. However, morula and blastocyst yield were improved (p < 0.05) after addition of EGF (30 ng/ml) as compared with maturation medium alone (29.3% vs 18.0%, respectively). Addition of r-hFSH to the maturation medium in the presence of 17β-estradiol (E2) significantly (p < 0.0001) increased the morula and blastocyst rate compared with maturation medium alone (40.3% vs 19.3%, respectively). The presence of BSA alone during IVM significantly reduced the developmental competence of oocytes as reflected by the morula and blastocyst yield. These results demonstrate the essential role of FSH, EGF and E2 on the kinetics of nuclear and cytoplasmic maturation that are essential for the formation of an egg capable of fertilisation and development. Also, supplementation of r-hFSH and E2 during IVM under our conditions increases morula and blastocyst yield following in vitro fertilisation and in vitro culture in defined medium. Finally, the presence of BSA as the only protein supplement during IVM may be detrimental to oocyte maturation.

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