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991.
Human follicle stimulating hormone is a pituitary glycoprotein that is essential for the maintenance of ovarian follicle development and testicular spermatogenesis. Like other members of the glycoprotein hormone family, it contains a common a subunit and a hormone specific subunit. Each subunit contains two glycosylation sites. The specific structures of the oligosaccharides of human follicle stimulating hormone have been shown to influence both thein vitro andin vivo bioactivity. Since the carbohydrate structure of a protein reflects the glycosylation apparatus of the host cells in which the protein is expressed, we examined the isoform profiles,in vitro bioactivity and metabolic clearance of a preparation of purified recombinant human follicle stimulating hormone derived from a stable, transfected Sp2/0 myeloma cell line, and pituitary human follicle stimulating hormone. Isoelectric focussing and chromatofocussing studies of human follicle stimulating hormone preparations both showed a more basic isoform profile for the recombinant human follicle stimulating hormone compared to that of pituitary human follicle stimulating hormone. The recombinant human follicle stimulating hormone had a significantly higher radioreceptor activity compared to that of pituitary human follicle stimulating hormone, consistent with a greaterin vitro potency. Pharmacokinetic studies in rats indicated a similar terminal half life (124 min) to that of the pituitary human follicle stimulating hormone (119 min). Preliminary carbohydrate analysis showed recombinant human follicle stimulating hormone to contain high mannose and/or hybrid type, in addition to complex type carbohydrate chains, terminating with both2,3 and2,6 linked sialic acids. These results demonstrate that recombinant human follicle stimulating hormone made in the Sp2/0 myeloma cells is sialylated, has a more basic isoform profile, and has a greaterin vitro biological potency compared to those of the pituitary human follicle stimulating hormone. 相似文献
992.
We designed an Integrated Media Preparation System (IMPS) for continuous, on-line preparation of cell culture media and delivery to intermediate storage vessels or directly to a bioreactor. Key components of the IMPS include: a high precision, continuous fluid mixing device; formulation-specific liquid medium concentrates; validated process controls and membrane filtration; and automated dispensing into large volume flexible plastic containers. The IMPS system is designed to produce sterile, single-strength liquid medium from common raw materials at a delivery rate of 1000–3000 liters per hour and will manufacture homogenous batches from several thousand liters to over 60,000 liters. Fortified nutrient media prepared from multi-component 50X concentrates have been demonstrated to accelerate bioreactor seed chains, increase product yield, and reduce the overall manufacturing cost of nutrient medium. A productivity matrix will analyze the fully-loaded costs and contrast alternative methods for media preparation against projected biological yield.Abbreviations IMPS
Integrated Media Preparation System
- 50X
Nutrient fluid components formulated at fifty-fold final use concentration
- 1X
Nutrient fluid formulated at final, single-strength use concentration
- cGMP
Current Good Manufacturing Practices
- SCADA
Supervisory Control and Data Acquisition
- PLC
Process Logic Controller
- LTI
Life Technologies, Inc.
- WFI
Water for Injection
- CIP
Clean in place
- SIP
Sterilize in place
- HPLC
High performance liquid chromatography
- DMEM
Dulbecco's Modified Eagle's Medium 相似文献
993.
Large scale production of recombinant mouse and rat growth hormone by fed-batch GS-NSO cell cultures
Zhou W Bibila T Glazomitsky K Montalyo J Chan C Distefano D Munshi S Robinson D Buckland B Aunins J 《Cytotechnology》1996,22(1-3):239-250
Investigations of biological effects of prolonged elevation of growth hormone in animals such as mice and rats require large amounts of mouse and rat growth hormone (GH) materials. As an alternative to scarce and expensive pituitary derived materials, both mouse and rat GH were expressed in NSO murine myeloma cells transfected with a vector containing the glutamine synthetase (GS) gene and two copies of mouse or rat GH cDNA. For optimal expression, the mouse GH vector also contained sequences for targeting integration by homologous recombination. Fed-batch culture processes for such clones were developed using a serum-free, glutamine-free medium and scaled up to 250 L production scale reactors. Concentrated solutions of proteins, amino acids and glucose were fed periodically to extend cell growth and culture lifetime, which led to an increase in the maximum viable cell concentration to 3.5×109 cells/L and an up to 10 fold increase in final mouse and rat rGH titers in comparison with batch cultures. For successful scale up, similar culture environmental conditions were maintained at different scales, and specific issues in large scale reactors such as balancing oxygen supply and carbon dioxide removal, were addressed. Very similar cell growth and protein productivity were obtained in the fed-batch cultures at different scales and in different production runs. The final mouse and rat rGH titers were approximately 580 and 240 mg/L, respectively. During fed-batch cultures, the cell growth stage transition was accompanied by a change in cellular metabolism. The specific glucose consumption rate decreased significantly after the transition from the growth to stationary stage, while lactate was produced in the exponential growth stage and became consumed in the stationary stage. This was roughly coincident with the beginning of ammonia and glutamate accumulation at the entry of cells into the stationary stage as the result of a reduced glutamine consumption and periodic nutrient additions. 相似文献
994.
The traditional sampling method for estimating frequency (the number of sub-quadrats containing a basal part of the organisms) is compared, using both computer simulations and direct comparison in the field, to two new methods that use a compound series of variable-sized concentric sub-quadrats. Both the new frequency-score and the new importance-score methods are closer approximations of density than is the standard frequency method, and the estimates produced by both of the new methods are less affected by the choice of sub-quadrat size and the spatial distribution (dispersion) of the organisms (i.e. clumping and regularity). Thus, the two nested-quadrat methods appear to ameliorate the usual frequency limitations associated with sub-quadrat size and organism dispersion, by the use of a range of different sub-quadrat sizes. This is important in community studies, where the component species may show a wide range of densities and dispersions. Both of the new methods are easily employed in the field. The importance-score method involves no more sampling effort than does standard qualitative (presence-absence) sampling, and it can therefore be used to sample a larger quadrat area than would normally be used for frequency sampling. This makes the method much more cost-effective as a means of estimating abundance, and it allows a greater number of the rarer species to be included in the sampling. The frequency-score method is more time-consuming, but it is capable of detecting more subtle community patterns. This means that it is particularly useful for the study of species-poor communities or where small variations in composition need to be detected. 相似文献
995.
The nervous system of Tricladida. I. Neuroanatomy ofProcerodes littoralis (Maricola,Procerodidae): An immunocytochemical study 总被引:3,自引:0,他引:3
Maria Reuter Margaretha K. S. Gustafsson Cecilia Sahlgren David W. Halton Aaron G. Maule Chris Shaw 《Invertebrate neuroscience : IN》1995,1(2):113-122
The organization of the nervous system ofProcerodes littoralis (Tricladida, Maricola, Procerodidae) was studied by immunocytochemistry, using antibodies to authentic flatworm neuropeptide
F (NPF) (Moniezia expansa). Compared to earlier investigations of the neuroanatomy of tricladid flatworms, the pattern of NPF immunoreactivity inProcerodes littoralis reveals differences in the following respects: 1. Shape and structure of the brain. 2. Number and composition of longitudinal
nerve cords. 3. Shape of branches of, and transverse connections between, main ventral nerve cords. 4. Composition of the
pharyngeal nervous system. The rich innervation by NPF immunoreactive (IR) fibres and cells of the subepithelial muscle layer,
the pharynx musculature and the musculature of the male copulatory apparatus indicates a neurotransmitter or neuromodulatory
influence on muscular activity. 相似文献
996.
The nervous system of Tricladida. II. Neuroanatomy ofDugesia tigrina (Paludicola,Dugesiidae): An immunocytochemical study 总被引:2,自引:0,他引:2
997.
David McElroy Douglas A. Chamberlain Eunpyo Moon Kate J. Wilson 《Molecular breeding : new strategies in plant improvement》1995,1(1):27-37
The use of reporter genes to characterise sequence elements that act to regulate gene expression in transgenic plants has been vital to the development of foreign gene expression strategies for use in cereal transformation. ThegusA locus ofEscherichia coli, which encodes the enzyme-glucuronidase (GUS), is by far the most popular reporter gene used in plant transformation. In this paper we extend the utility of the GUS reporter gene system in cereal transformation by describing and evaluating a number of novel constructs suitable for use in direct gene transfer experiments. These plasmids are all available from the Molecular Genetic Resource Service of the Center for the Application of Molecular Biology to International Agriculture. 相似文献
998.
Ariela Baruch Moshe Shani David R. Hurwitz Itamar Barash 《Genesis (New York, N.Y. : 2000)》1995,16(3):241-252
999.
The extent of contamination of soils by toxic heavy metals not only depends on the rate of loading of the metal but also on the nature of the adsorbing surfaces, the degree of alkalinity or acidity of the soil and the presence of aqueous complexant ligands. This work reports on the role of pH on the retention of Cd, Hg, Pb and Zn by two soils and on the influence of the chloride, Cl‐, ion on the chemical speciation and retention of the four metals. Batch adsorption experiments were conducted from pH 3 to 7 in the presence of either 0.1 M LiCl or LiClO4. The results of the study showed that high concentrations of Cl‐ ions can greatly decrease the retention of Hg and have an increasingly lesser effect on Cd, Pb and Zn retention. The effect of the Cl‐ons was directly related to the metal‐Cl formation constants. The results of computer modeling of Cd and Hg retention by goethite and humic acid fractions indicated the relative importance of aqueous vs. surface complexation on metal retention. For organic surfaces, which do not form ternary surface complexes, the presence of aqueous complexant ligands should always decrease the adsorption of the metal. For mineral surfaces, which do form ternary surface complexes, there may be increased or decreased metal retention depending on the formation constant of the aqueous metal‐ligand species, the intrinsic complexation constants for the various binary and ternary complexes of the metal and the concentration of the complexant ligand. Thus for Hg, which forms very strong aqueous species with Cl‐ ions, reduced adsorption on goethite was predicted in the presence of 0.1 M LiCl, while enhanced adsorption was predicted for Cd and Pb. The results suggest caution in the disposal of Cl‐containing wastes onto metal‐contaminated soils. The deleterious effects of Cl‐ ion addition would be greatest for soils with relatively high organic matter contents and low contents of hydrous ferric oxides. 相似文献
1000.
Dynein and kinesin have been implicated as the molecular motors that are responsible for the fast transport of axonal membranous organelles and vesicles. Experiments performed in vitro with partially reconstituted preparations have led to the hypothesis that kinesin moves organelles in the anterograde direction and dynein moves them in the retrograde direction. However, the molecular basis of transport directionality remains unclear. In the experiments described here, carboxylated fluorescent beads were injected into living Mauthner axons of lamprey and the beads were observed to move in both the anterograde and retrograde directions. The bead movement in both directions required intact microtubules, occurred at velocities approaching organelle fast transport in vivo, and was inhibited by vanadate at concentrations that inhibit organelle fast transport. When living axons were injected with micromolar concentrations of vanadate and irradiated at 365 nm prior to bead injections, a treatment that results in the V1 photolysis of dynein, the retrograde movement of the beads was specifically abolished. Neither the ultraviolet irradiation alone nor the vanadate alone produced the retrograde-specific inhibition. These results support the hypothesis that dynein is required for retrograde, but not anterograde, transport in vivo. © 1995 John Wiley & Sons, Inc. 相似文献