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  • Lactic acid production through cell-recycle repeated-batch bioreactor. 12721439

    The effect of various nitrogen sources on cell growth and lactic acid production was investigated. The most effective nitrogen source was yeast extract; more yeast extract gave higher cell growth and lactic acid productivity. Yeast extract dosage and cell growth were proportional up to a yeast extract concentration of 30 g/L, and lactic acid productivity was linearly correlated up to a yeast extract dosage of 25 g/L. However, increasing the yeast extract content raises the total production cost of lactic acid. Therefore, we attempted to find the optimum yeast extract dosage for a repeated-batch operation with cell recycling. The results show that when using Enterococcus faecalis RKY1 only 26% of the yeast extract dosage for a conventional batch fermentation was sufficient to produce the same amount of lactic acid, whereas the lactic acid concentration in the product stream (92-94 g/L) and lactic acid productivity (6.03-6.20 g/[L x h]) were similar to those of a batch operation. Furthermore, long-term stability was established.
    Tipo de documento:
    Referencia
    Referencia del producto:
    20-108
    Nombre del producto:
    Assay Dilution Buffer I (ADBI)

  • Cyclosporine-A treatment inhibits the expression of metabotropic glutamate receptors in rat thymus. 12666991

    We have evaluated the expression of metabotropic glutamate receptors (mGluR subtypes 2/3, 4 and 5) in rat thymus under normal and experimental conditions after 2 and 21 days of cyclosporine-A treatment. In normal rats, immunohistochemical analysis showed that expression of mGluRs was high in dendritic cells and lymphocytes of the medulla whereas it was weak in lymphocytes of the cortex. However, there were some differences in the expression of mGluRs subtypes. mGluR5 showed strong expression in lymphocytes of medulla and dendritic cells. mGluR2/3 and mGluR4 were moderately expressed in lymphocytes and dendritic cells of the medulla and weakly in cortical lymphocytes. Immunoblotting showed moderate levels of mGluR2/3 and mGluR4 and strong levels of mGluRS. After 2 days of cyclosporine-A treatment, we observed by immunohistochemistry and immunoblotting a distinct decrease in all mGluRs and their expression had almost completely disappeared after 21 days of treatment. The results clearly indicate that: 1) mGluR2/3, 4 and 5 are widely expressed in thymic cells; 2) the mGluR5 subtype is expressed most strongly in medullary cells; and 3) cyclosporine-A rapidly inhibits expression of all mGluR subtypes after 2 days of treatment and their complete disappearance after prolonged treatment. These findings may indicate a possible mechanism by which cyclosporine-A produces its immunosupressive effects.
    Tipo de documento:
    Referencia
    Referencia del producto:
    03-105

  • Permeation associated with three-phase-partitioning method on release of green fluorescent protein. 12721429

    Transformed cells of Escherichia coli expressing recombinant green fluorescent protein (GFPuv) were subjected to two methods of extraction: (1) freezing/thawing/sonication (FTS) cycles prior to the three-phase partitioning (TPP) method, or (2) directly to TPP extraction. The amount of GFPuv released by the FTS plus TPP method varied: 374 microg/mL (first cycle), 93-442 microg/mL (second cycle), 32-359 microg/mL (third cycle), 18-115 microg/mL (fourth cycle). The GFPuv yields by the second method (TPP only) were, 23-54 microg/mL for the first extract and 33-91 microg/mL for the second. The FTS plus TPP method released similar amounts of GFPuv to that extracted by TPP; and provided a better mixture elution through the hydrophobic interaction column: 13-63 microg/mL for FTS plus TPP methods, and 2.5-13 microg/mL for TPP. The results showed that although selective permeation is a more laborious methodology, it was more efficient for obtaining of GFPuv in relation to the direct extraction of the cells for TPP.
    Tipo de documento:
    Referencia
    Referencia del producto:
    20-108
    Nombre del producto:
    Assay Dilution Buffer I (ADBI)

  • Sugar monomer and oligomer solubility: data and predictions for application to biomass hydrolysis. 12721484

    Oligomer solubility could potentially play an important role in controlling the rates and yields in the thermochemical hydrolysis of hemicellulose as a pretreatment for subsequent enzymatic conversion of cellulose. However, limited data or models are available to describe the aqueous solubility of sugar monomers and oligomers. In this work, we measured the solubilities of sugars common to many biomass feedstocks in the temperature range of 25-30 degrees C. Then we reviewed solubility models for sugars from the open literature. Finally, we applied models to test their ability to describe this and other data reported in the literature. It was found that the solubility of sugar monomers was not well described by the ideal solubility law or other more complex models. However, with an empirical adjustment to the enthalpy of fusion, the ideal solubility law was able to approximately predict the solubility of cello-oligomers. Based on these results, solubilities for low molecular weight xylo-oligomers are predicted to investigate their possible importance in pretreatment and define further experimental measurements needed to improve our understanding of sugar and oligomer solubility.
    Tipo de documento:
    Referencia
    Referencia del producto:
    20-108
    Nombre del producto:
    Assay Dilution Buffer I (ADBI)

  • Immunolocalization of alphaV, alpha3 and beta1 integrins in the human placenta with pre-eclampsia. 13677619

    Signs of pre-eclampsia are considered to be caused by maternal endothelial dysfunction due to circulating factors of placental origin. Integrins are a large family of cell surface, proteins that serve as receptors involved in cell-cell and cell-matrix interactions during placentation. Therefore, low expression of integrins or the lack of it may be encountered during pre-eclampsia. In the present study, we investigated the immunolocalisation of integrins alphaV, alpha3 and beta1 in placentas of normal and pre-eclamptic women. Thirty-two placentas from pre-eclamptic (n = 14) and normotensive (n = 18) women were used. Immunohistochemical staining was performed on formalin-fixed, paraffin-embedded tissue specimens, using anti-alphaV, anti-alpha3 and anti-beta1 antibodies and the indirect immunoperoxidase technique. A semi-quantitative grading system (HSCORE) was used to compare immunohistochemical staining intensities. Distribution patterns of alphaV, alpha3 and beta1 integrins were detected in cytotrophoblasts and Hofbauer cells in normal and pre-eclamptic placentas. Immunostaining of alphaV and beta1 integrins was slightly decreased in pre-eclamptic samples but alpha3 integrin immunostaining was similar in pre-eclamptic and normal placentas. Decreased immunostaining of integrins in the cytotrophoblasts may considered to be a structural basis for decreased placental perfusion in pre-eclampsia.
    Tipo de documento:
    Referencia
    Referencia del producto:
    03-105

  • Characterization of polysaccharide production of haemophilus influenzae Type b and its relationship to bacterial cell growth. 14515024

    Haemophilus influenzae type b (Hib) causes invasive infections in infants and young children. Vaccines consisting of Hib capsular polysaccharide (polymer of ribosylribitol phosphate [PRP]) conjugated to a protein are effective in the prevention of such infections. The production of capsular polysaccharide type b was studied in three cultivation conditions: single, glucose pulse, and repeated batch. Specific polysaccharide production (Yp/x) was calculated for all experiments, showing the following values: 67 (single-batch cultivation), 71 (glucose pulse), 75 (repeated-batch cultivation, first batch), and 87 mg of PRP/g of dry cell weight (DCW) (repeated-batch cultivation, second batch). Biomass concentration reached approximately 1.8 g of DCW/L, while polysaccharide concentration was about approximately 132 mg/L in the three fermentation runs. Polysaccharide synthesis is associated with cell growth in all studied conditions as established by Kono's analysis and Luedeking-Piret's model.
    Tipo de documento:
    Referencia
    Referencia del producto:
    03-110
    Nombre del producto:
    RIPAb+ Ago2 - RIP Validated Antibody and Primer Set

  • Immobilization of glutaryl-7-aminocephalosporanic acid acylase on silica gel and enhancement of its stability. 12665670

    Glutaryl-7-aminocephalosporanic acid (GL-7-ACA) acylase is an enzyme that converts GL-7-ACA to 7-aminocephalosporanic acid, a starting material for semisynthetic cephalosporin antibiotics. In this study, optimal conditions for the immobilization of GL-7-ACA acylase were determined by experimental observations and statistical methods. The optimal conditions were as follows: 1.1 M phosphate buffer (pH 8.3) as buffer solution, immobilization temperature of 20 degrees C, and immobilization time of 120 min. Unreacted aldehyde groups were quenched by reaction with a low-molecular-weight material such as L-lysine, glycine, and ethanolamine after immobilization in order to enhance the activity of immobilized GL-7-ACA acylase. The activities of immobilized GL-7-ACA acylase obtained by using the low-molecular-weight materials were higher than those obtained by immobilized GL-7-ACA acylase not treated with low-molecular-weight materials. In particular, the highest activity of immobilized GL-7-ACA acylase was obtained using 0.4% (v/v) ethanolamine. We also investigated the effect of sodium cyanoborohydride in order to increase the stability of the linkage between the enzyme and the support. The effect on operational stability was obvious: the activity of immobilized GL-7-ACA acylase treated with 4% (w/w) sodium cyanoborohydride remained almost 100% after 20 times of reuse.
    Tipo de documento:
    Referencia
    Referencia del producto:
    03-104
    Nombre del producto:
    RIPAb+ CUGBP1 - RIP Validated Antibody and Primer Set

  • Expression of transcription factors and matrix genes in response to serum stimulus in vascular smooth muscle cells. 12691261

    During atherogenesis vascular smooth muscle cells are converted from a contractile into a synthetic phenotype characterized by enhanced matrix production. The transcription factors Gax and GATA-6 are considered negative, and Oct-1 positive regulators of the synthetic phenotype. Since the phenotype transition can be induced by culturing the cells with serum, we followed the expression of Gax, GATA-6 and Oct-1, integrins and matrix genes in quiescent porcine vascular smooth muscle cells after serum application. Comparisons were made between enzymatically released primary smooth muscle cells and cells grown out from explants of the medial layer of porcine aorta. The serum-mediated down-regulation of Gax was more intense than that of GATA-6, and stronger in explant-derived than in primary cells. Serum was without influence on the expression of Oct-1. Changes in the expression of the transcription factors preceded the induction of integrin alpha2 and the down-regulation of decorin, while mRNAs for laminin beta1 and osteopontin rose immediately after serum stimulation. Primary cells reacted more rapidly than explant cells with respect to changes in laminin isoforms. Studies with a Gax-expressing adenovirus indicated that among all the gene products tested only the expression of integrin alpha2 responded to Gax induction. Thus, our data show that i) Gax should be considered a transcription factor being directly responsible for only few aspects of the phenotypic conversion of smooth muscle cells and that ii) explant cells may represent a subpopulation of smooth muscle cells, which differ from the total population of smooth muscle cells, as obtained in primary culture, in their response to serum stimuli.
    Tipo de documento:
    Referencia
    Referencia del producto:
    03-119
    Nombre del producto:
    RIPAb+™ CUGBP2 - RIP Validated Antibody and Primer Set

  • Cells previously identified as retinal stem cells are pigmented ciliary epithelial cells. 19346468

    It was previously reported that the ciliary epithelium (CE) of the mammalian eye contains a rare population of cells that could produce clonogenic self-renewing pigmented spheres in culture. Based on their ability to up-regulate genes found in retinal neurons, it was concluded that these sphere-forming cells were retinal stem cells. This conclusion raised the possibility that CE-derived retinal stem cells could help to restore vision in the millions of people worldwide who suffer from blindness associated with retinal degeneration. We report here that human and mouse CE-derived spheres are made up of proliferating pigmented ciliary epithelial cells rather than retinal stem cells. All of the cells in the CE-derived spheres, including the proliferating cells, had molecular, cellular, and morphological features of differentiated pigmented CE cells. These differentiated cells ectopically expressed nestin when exposed to growth factors and low levels of pan-neuronal markers such as beta-III-tubulin. Although the cells aberrantly expressed neuronal markers, they retained their pigmented CE cell morphology and failed to differentiate into retinal neurons in vitro or in vivo. Our results provide an example of a differentiated cell type that can form clonogenic spheres in culture, self-renew, express progenitor cell markers, and initiate neuronal differentiation that is not a stem or progenitor cell. More importantly, our findings highlight the importance of shifting the focus away from studies on CE-derived spheres for cell-based therapies to restore vision in the degenerating retina and improving techniques for using ES cells or retinal precursor cells.
    Tipo de documento:
    Referencia
    Referencia del producto:
    AB5585

  • Lesioning noradrenergic neurons of the locus coeruleus in C57Bl/6 mice with unilateral 6-hydroxydopamine injection, to assess molecular, electrophysiological and biochemi ... 22542679

    The locus coeruleus (LC) is the major loci of noradrenergic innervation to the forebrain. Due to the extensive central nervous system innervation of the LC noradrenergic system, a reduction in the number of LC neurons could result in significant changes in noradrenergic function in many forebrain regions. LC noradrenergic neurons were lesioned in adult male C57Bl/6 mice with the unilateral administration of 6-hydroxydopamine (6OHDA) (vehicle on the alternate side). Noradrenergic markers were measured 3 weeks later to determine the consequence of LC loss in the forebrain. Direct administration of 6OHDA into the LC results in the specific reduction of noradrenergic neurons in the LC (as measured by electrophysiology, immunoreactivity and in situ hybridization), the lateral tegmental neurons and dopaminergic neurons in the substantia nigra (SN) and ventral tegmental region were unaffected. The loss of LC noradrenergic neurons did not result in compensatory changes in the expression of mRNA for norepinephrine (NE)-synthesizing enzymes. The loss of LC noradrenergic neurons is associated with reduced NE tissue concentration and NE transporter (NET) binding sites in the frontal cortex and hippocampus, as well as other forebrain regions such as the amygdala and SN. Adrenoreceptor (AR) binding sites (α(1)- and α(2)-AR) were not significantly affected on the 6OHDA-treated side compared to the vehicle-treated side, although there is a reduction of AR binding sites on both the vehicle- and 6OHDA-treated side in specific forebrain regions. These studies indicate that unilateral stereotaxic injection of 6OHDA into mice reduces noradrenergic LC neurons and reduces noradrenergic innervation to many forebrain regions, including the contralateral side.
    Tipo de documento:
    Referencia
    Referencia del producto:
    AB152
    Nombre del producto:
    Anti-Tyrosine Hydroxylase Antibody