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  • Effects of ileal interposition on glucose metabolism in obese rats with diabetes. 22316438

    Ileal interposition (IT), in which the distal ileum is transposed isoperistaltically into the proximal jejunum, is considered as a procedure for metabolic or antidiabetes surgery. Our aim was to study the effects of IT on glycemic control, fat metabolism, and hormonal changes in obese rats with spontaneous diabetes.
    Document Type:
    Reference
    Product Catalog Number:
    EZRMGIP-55K
    Product Catalog Name:
    Rat/Mouse GIP (total) ELISA - (Rat/Mouse GIP (total) ELISA)

  • Application of multiplex PCR for characterization of human embryonic stem cells (hESCs) and its differentiated progenies. 20530724

    Techniques to evaluate gene expression profiling, including real-time quantitative PCR, TaqMan low-density arrays, and sufficiently sensitive cDNA microarrays, are efficient methods for monitoring human embryonic stem cell (hESC) cultures. However, most of these high-throughput tests have a limited use due to high cost, extended turnaround time, and the involvement of highly specialized technical expertise. Hence, there is a paucity of rapid, cost-effective, robust, yet sensitive methods for routine screening of hESCs. A critical requirement in hESC cultures is to maintain a uniform undifferentiated state and to determine their differentiation capacity by showing the expression of gene markers representing all germ layers, including ecto-, meso-, and endoderm. To quantify the modulation of gene expression in hESCs during their propagation, expansion, and differentiation via embryoid body (EB) formation, the authors developed a simple, rapid, inexpensive, and definitive multimarker, semiquantitative multiplex RT-PCR (mxPCR) platform technology. Among the 15 gene primers tested, 4 were pluripotent markers comprising set 1, and 3 lineage-specific markers from each ecto-, meso-, and endoderm layers were combined as sets 2 to 4, respectively. The authors found that these 4 sets were not only effective in determining the relative differentiation in hESCs, but were easily reproducible. In this study, they used the HUES-7 cell line to standardize the technique, which was subsequently validated with HUES-9, NTERA-2, and mouse embryonic fibroblast cells. This single-reaction mxPCR assay was flexible and, by selecting appropriate reporter genes, can be designed for characterization of different hESC lines during routine maintenance and directed differentiation.
    Document Type:
    Reference
    Product Catalog Number:
    Multiple
    Product Catalog Name:
    Multiple

  • Ghrelin vaccination decreases plasma MCP-1 level in LDLR(-/-)-mice. 19751783

    Ghrelin is a novel peptide hormone having growth hormone releasing activity and many endocrine and metabolic functions. In rats and pigs, ghrelin immunizations have recently been shown to induce an antibody response against ghrelin simultaneously with a decrease in body weight gain. Our aim was to test the role of ghrelin immunization on atherosclerosis and weight gain in mice. LDLR(-/-)-mice (n=36) were immunized with ghrelin-PADRE, PADRE alone and PBS and then placed on a high fat diet for 22 weeks. Weight gain and food intake were followed throughout the study. Acylated and total ghrelin, cytokines and MCP-1 were analyzed from plasma using commercial kits. Stomach ghrelin was assessed using qRT-PCR and immunohistochemistry. Atherosclerosis was determined from aorta and cross-sections at the end of study. Mice immunized with ghrelin-PADRE developed high plasma IgG titers to ghrelin simultaneously with a significant increase in plasma acylated and total ghrelin levels. Plasma MCP-1 levels decreased in mice immunized with ghrelin-PADRE compared to mice immunized with PADRE and PBS. There were no differences in atherosclerosis determined from aorta and cross-sections as well as in body weights and food intake in LDLR(-/-)-mice between the different immunization groups. Our data indicates that ghrelin-PADRE vaccination induces a strong exclusive IgG response to ghrelin and increases plasma acylated and total ghrelin levels in mice. Ghrelin vaccination decreases plasma MCP-1 levels even though no effects on developing signs of atherosclerosis or weight gain in mice were observed.
    Document Type:
    Reference
    Product Catalog Number:
    EZRMI-13K
    Product Catalog Name:
    Rat/Mouse Insulin ELISA - (Rat/Mouse Insulin ELISA)

  • Role of the non-canonical notch ligand delta-like protein 1 in hormone-producing cells of the adult male mouse pituitary. 21756269

    To better understand the role of the non-canonical Notch ligand delta-like protein 1 (DLK1), in hormone-producing cells, we studied the cell distribution and subcellular localisation of DLK1 in the pituitary of male adult 129/SvJ mice, and analysed the variations in the hormone-producing cells associated with the lack of this gene in Dlk1 knockout mice. The results obtained showed the presence of DLK1-immunoreactive (ir) cells in all hormone-producing cells of the anterior pituitary. Immunoelectron microscopy showed DLK1-ir in the rough endoplasmic reticulum and inside secretory vesicles, suggesting that DLK1 is released together with pituitary hormones. Moreover, we found that prolactin (PRL)-DLK1-ir cells are in intimate contact with follicle-stimulating hormone (FSH)-ir-DLK1-negative cells. In Dlk1 knockout mice, we detected a significantly lower number of gowth hormone (GH)-ir cells, a reduction in the FSH and PRL immunostaining intensity, and a significant decrease in FSH mRNA expression compared to wild-type mice. An increase in pituitary GH mRNA expression and serum leptin levels was also found. These findings provide evidence supporting several regulatory functions of DLK1 in the pituitary gland.
    Document Type:
    Reference
    Product Catalog Number:
    AB940

  • Detection of immunoglobulin isotypes from dried blood spots. 24333851

    The study was designed to determine the sensitivity and reproducibility of recovering immunoglobulin (Ig) isotypes (IgG subclasses, IgA, IgE and IgM classes) from dried blood spots (DBS), a methodologic subcomponent of the Upstate KIDS Study. A multiplexed Luminex assay was used for IgG1/2/3/4, IgA and IgM analysis; an ELISA was used for IgE. Plasma samples from de-identified patients were used to compare the Luminex assay with nephelometry, which is routinely used to quantify IgA, IgG and IgM in clinical samples. The IgE ELISA was compared to an immunofluorescence assay. Prior to evaluation of punches from newborn dried blood spots (NDBSs), recoveries of Ig from punches of cord blood DBSs (CBDBSs) vs. plasma from the same cord bloods were compared. Although the recoveries of Ig from plasma and DBSs were not comparable, which could be due to cell lysates in the DBS samples, the analyses were reproducible. Additionally, the levels of IgA, IgG2, IgG4, and IgM recovered from CBDBSs positively correlated with those in plasma. The DBS data is a relative value since it is not equivalent to the plasma concentration. The majority of Ig concentrations recovered from 108 newborns of the Upstate KIDs Study were within the range of newborn plasma Ig levels with the exception of IgG3. The IgG4 values displayed the greatest variance with a wide range (0.01-319 mg/dl), whereas, IgG1 values had the narrowest range (85.2-960.4 mg/dl).
    Document Type:
    Reference
    Product Catalog Number:
    HGAMMAG-301K
    Product Catalog Name:
    MILLIPLEX® MAP Human Isotyping Magnetic Bead Panel - Isotyping Multiplex Assay - (MILLIPLEX® MAP Human Isotyping Magnetic Bead Panel - Isotyping Multiplex Assay)

  • Mesenchymal stem cells secrete multiple cytokines that promote angiogenesis and have contrasting effects on chemotaxis and apoptosis. 22558198

    We have previously shown that mesenchymal stem cells (MSC) improve function upon integration in ischemic myocardium. We examined whether specific cytokines and growth factors produced by MSCs are able to affect angiogenesis, cellular migration and apoptosis. Conditioned media (CM) was prepared by culturing MSC for 48 hours. CM displayed significantly elevated levels of VEGF, Monocyte Chemoattractant Protein-1 (MCP-1), macrophage inflammatory protein-1α (MIP-1α), MIP-1β and monokine induced by IFN-γ (MIG) compared to control media. MSC contained RNA for these factors as detected by RT-PCR. CM was able to induce angiogenesis in canine vascular endothelial cells. MCP-1 and MIP-1α increased cell migration of MSC while VEGF reduced it. H9c2 cells treated with CM under hypoxic conditions for 24 hours displayed a 16% reduction in caspase-3 activity compared to controls. PI 3-kinase γ inhibitor had no effect on controls but reversed the effect of CM on caspase-3 activity. MCP-1 alone mimicked the protective effect of CM while the PI 3-Kγ inhibitor did not reverse the effect of MCP-1. CM reduced phospho-BAD (Ser112) and phospho-Akt (Ser473) while increasing phospho-Akt (Thr308). MCP-1 reduced the level of phospho-Akt (Ser473) while having no effect on the other two; the PI 3-Kγ inhibitor did not alter the MCP-1 effect. ERK 1/2 phosphorylation was reduced in CM treated H9c2 cells, and inhibition of ERK 1/2 reduced the phosphorylation of Akt (Ser473), Akt (Thr308) and Bad (Ser112). In conclusion, MSC synthesize and secrete multiple paracrine factors that are able to affect MSC migration, promote angiogenesis and reduce apoptosis. While both MCP-1 and PI3-kinase are involved in the protective effect, they are independent of each other. It is likely that multiple pro-survival factors in addition to MCP-1 are secreted by MSC which act on divergent intracellular signaling pathways.
    Document Type:
    Reference
    Product Catalog Number:
    ECM630
    Product Catalog Name:
    Fibrin In Vitro Angiogenesis Assay - (Fibrin In Vitro Angiogenesis Assay)

  • Robust, high-throughput solution for blood group genotyping. 20560530

    With the concomitant increase of blood transfusions and safety rules, there is a growing need to integrate high-throughput and multiparametric assays within blood qualification centers. Using a robust and automated solution, we describe a new method for extended blood group genotyping (HiFi-Blood 96) bringing together the throughput possibilities of complete automation and the microarray multiplexed analysis potential. Our approach provides a useful resource for upgrading blood qualification center facilities. A set of six single-nucleotide polymorphisms (SNPs) associated with clinically important blood group antigens (Kell, Kidd, Duffy, and MNS systems) were selected and the corresponding genotyping assays developed. A panel of 293 blood samples was used to validate the approach. The resulting genotypes were compared to phenotypes previously determined by standard serologic techniques, and excellent correlations were found for five SNPs out of six. For the Kell, Kidd, Duffy, and MNS3/MNS4 systems, high matching percentages of 100%, 98.9%, 97.7%, and 97.4% were obtained, respectively, whereas a concordance percentage of 83.3% only was attained for the MNS1/MNS2 polymorphism.
    Document Type:
    Reference
    Product Catalog Number:
    MSPLRS096

  • The medicinal plant goldenseal is a natural LDL-lowering agent with multiple bioactive components and new action mechanisms. 16885565

    Our previous studies have identified berberine (BBR), an alkaloid isolated from the Chinese herb huanglian, as a unique cholesterol-lowering drug that upregulates hepatic low density lipoprotein receptor (LDLR) expression through a mechanism of mRNA stabilization. Here, we demonstrate that the root extract of goldenseal, a BBR-containing medicinal plant, is highly effective in upregulation of liver LDLR expression in HepG2 cells and in reducing plasma cholesterol and low density lipoprotein cholesterol (LDL-c) in hyperlipidemic hamsters, with greater activities than the pure compound BBR. By conducting bioassay-driven semipurifications, we demonstrate that the higher potency of goldenseal is achieved through concerted actions of multiple bioactive compounds in addition to BBR. We identify canadine (CND) and two other constituents of goldenseal as new upregulators of LDLR expression. We further show that the activity of BBR on LDLR expression is attenuated by multiple drug resistance-1 (MDR1)-mediated efflux from liver cells, whereas CND is resistant to MDR1. This finding defines a molecular mechanism for the higher activity of CND than BBR. We also provide substantial evidence to show that goldenseal contains natural MDR1 antagonist(s) that accentuate the upregulatory effect of BBR on LDLR mRNA expression. These new findings identify goldenseal as a natural LDL-c-lowering agent, and our studies provide a molecular basis for the mechanisms of action.
    Document Type:
    Reference
    Product Catalog Number:
    ECM910
    Product Catalog Name:
    MDR1 Efflux Assay - (MDR1 Efflux Assay)

  • Color-coded fluorescence imaging of tumor-host interactions. 17406326

    Fluorescent proteins have the properties of being very bright with high quantum yield and are available in many colors. Tumor-host models consist of transgenic mice expressing green fluorescent protein (GFP) in essentially all cells and tissues or expressing GFP selectively in specific tissues such as blood vessels. Particularly useful are the corresponding nude mice transgenic for GFP expression, as they can accept human tumors. When tumor cells expressing red fluorescent protein are implanted in mice expressing GFP, various types of tumor-host interactions can be observed, including those involving host blood vessels, lymphocytes, tumor-associated fibroblasts, macrophages, dendritic cells and others. The 'color-coded' tumor-host models enable imaging and therefore a deeper understanding of the host cells involved and their function in tumor progression. Approximately 4-8 weeks are needed for these procedures.
    Document Type:
    Reference
    Product Catalog Number:
    CBL1337
    Product Catalog Name:
    Anti-PECAM-1 Antibody, clone 390 - (Anti-PECAM-1 Antibody, clone 390)

  • Crude subcellular fractionation of cultured mammalian cell lines. 20003239

    The expression and study of recombinant proteins in mammalian culture systems can be complicated during the cell lysis procedure by contaminating proteins from cellular compartments distinct from those within which the protein of interest resides and also by solubility issues that may arise from the use of a single lysis buffer. Partial subcellular fractionation using buffers of increasing stringency, rather than whole cell lysis is one way in which to avoid or reduce this contamination and ensure complete recovery of the target protein. Currently published protocols involve time consuming centrifugation steps which may require expensive equipment and commercially available kits can be prohibitively expensive when handling large or multiple samples.
    Document Type:
    Reference
    Product Catalog Number:
    05-516
    Product Catalog Name:
    Anti-Ras Antibody, clone RAS10 - (Anti-Ras Antibody, clone RAS10)