Sclerostin Antibody | AbD09097_h/mIgG2a

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Sclerostin Antibody | AbD09097_h/mIgG2a gallery image 1

Published customer image:
HuCAL anti Human sclerostin antibody, clone AbD09097 (HCA230Z) used in a functional assay to inhibit internalization of sclerostin and sclerostin modulated LRP6 signaling.
Image caption:
Inhibition of Sclerostin function by antibodies.
(A) Sclerostin-GFP was pre-incubated with Fc or the different Fabs raised against Sclerostin for 30 minutes at room temperature, next the mixtures were added to 293-LRP6 cells for 1.5h at 4°C. After binding the cells were transferred to a 37°C incubator for 30 minutes, next the cells were fixed and confocal images were taken. (B) KS483 cells were affinity labeled with [125I]Sclerostin that was pre-incubated for 30 minutes at room temperature with controls (Fc and aGFP-IgG) or Sclerostin neutralizing antibodies (AbD9097 and aSCL-IgG), followed by chemical crosslinking. Cells were lysed and samples were separated by SDS-PAGE. The radioactive signal was visualized using a phosphorimager screen. (C) KS483 cells were transiently transfected with the BAT-luc reporter construct. The cells were stimulated with conditioned control media (pcDNA CM) or Wnt3a conditioned media (Wnt3a CM), together or without Sclerostin conditioned (SCL CM) media that was pre-incubated with either anti GFP-IgG2a (aGFP-IgG) or anti Sclerostin-IgG2a (aSCL-IgG) for 30 minutes at room temperature. (D) KS483 cells were stimulated with control conditioned media or Wnt3a conditioned media (Wnt3a CM), together or without Sclerostin conditioned media that was pre-incubated with either anti GFP-IgG2a (aGFP-IgG) or anti Sclerostin-IgG2a (aSCL-IgG) for 30 minutes at room temperature. RNA was isolated and AXIN2 mRNA levels were determined by qRT-PCR. (E) KS483 cells were stimulated with the indicated conditioned media, which were pre-incubated with the indicated Fab antibodies. Three days after stimulation cells were fixed and stained for alkaline phosphatase activity. Results were quantified using spectrometry (F).

From: van Dinther M, Zhang J, Weidauer SE, Boschert V, Muth E-M, Knappik A, et al. (2013)
Anti-Sclerostin Antibody Inhibits Internalization of Sclerostin and Sclerostin-Mediated Antagonism of Wnt/LRP6 Signaling.
PLoS ONE 8(4): e62295.

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  • Sclerostin Antibody | AbD09097_h/mIgG2a thumbnail image 1
  • Human anti Sclerostin:Preservative Free
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  • Product Type
    Monoclonal Antibody
  • Clone
    AbD09097_h/mIgG2a
  • Isotype
    IgG2a
1 Formats Available
    Product CodeApplicationsDatasheetMSDSPack SizeList PriceQuantity
    HCA230ZC, FNdatasheet pdfdatasheet pdf0.1 mg
    HCA230Z
    Summary
    Secondary Antibodies
    Negative Isotype Controls
    Useful Reagents
    Positive Controls
    Histology Controls
    More Images
    References
    Reviews
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    • Human anti sclerostin, clone AbD09097_h/mIgG2a specifically recognises human and mouse sclerostin, also known as SOST. Sclerostin is a secreted extracellular matrix protein that is expressed at low levels in bone, bone marrow and cartilage. It may also be detected in other tissues such as kidney and liver. Sclerostin is important in the negative regulation of bone growth with mutations in the sclerostin gene resulting in conditions associated with high bone mass such as sclerosteosis and van Buchem disease. Sclerostin research may facilitate the development of future treatments for diseases associated with bone loss such as osteoporosis (Papapoulos 2011).

      Sclerostin mediates its inhibitory effect on bone formation by directly blocking the Wnt signaling pathway. In addition, it has been reported that sclerostin may associate with intracellular bone morphogenic protein 7 (BMP7), and block its secretion in osteocytes (Krause et al. 2010).

      Clone AbD09097_h/mIgG2a has been demonstrated to block the binding of sclerostin to the Wnt co-receptors, low density lipoprotein receptor-related protein (LRP) 5 and LRP6 on KS483 cells. In a BAT-luc reporter assay, clone AbD09097_h/mIgG2a showed a significant rescue of the inhibition by Sclerostin on the Wnt3a induced BAT-luc activity. Clone AbD09097_h/mIgG2a was also able to rescue the inhibition by sclerostin on Wnt3a-induced AXIN2 mRNA expression and the sclerostin-mediated inhibition of alkaline phosphatase activity induced by Wnt3a (van Dinther et al. 2013).

      Clone AbD09097_h/mIgG2a is a fully human/mouse chimeric antibody, developed from the inhibitory Fab antibody AbD09097(van Dinther et al. 2013).
    • Intended Use
    • Target Species
      Human
    • Species Cross-Reactivity
      Target SpeciesCross Reactivity
      Mouseyes
      N.B. Antibody reactivity and working conditions may vary between species.
    • Product Form
      Chimeric human/mouse IgG2a antibody selected from the HuCAL® phage display library and expressed in a human cell line. The antibody has a Fab portion of human origin and an Fc portion from mouse IgG2a. It can be detected by either anti-human Fab-specific or by anti-mouse Fc specific secondary antibodies. This antibody is supplied as a liquid.
    • Reconstitution
    • Preparation
      Purified IgG prepared by affinity chromatography on Protein A
    • Preservative Stabilisers
      None present
    • Immunogen
      Recombinant human and murine sclerostin.
    • Purity
    • Affinity
      The monovalent intrinsic affinity of AbD09097_h/mIgG2a was measured as Kd = 253 nM to human sclerostin and Kd = 49 nM to mouse sclerostin. Affinity measurements were performed using the ProteOn device (Bio-Rad) on recombinant sclerostin immobilized on an activated ProteOn GLC sensor chip.
    • Approx. Protein Concentrations
      Total protein concentration 0.5 mg/ml
    • Reagents In The Kit
    • Preparing The Antibody
    • Test Principle
    • Buffer Solution
      Phosphate buffered saline
    • Storage
      Store at -20oC only.
      Storage in frost-free freezers is not recommended.
      This product should be stored undiluted. Avoid repeated freezing and thawing as this may denature the antibody.
    • Shelf Life
      12 months from date of despatch
    • GO Terms
      negative regulation of Wnt receptor signaling pathway involved in dorsal/ventral axis specification
      negative regulation of canonical Wnt receptor signaling pathway
      negative regulation of BMP signaling pathway
      protein binding
      Wnt receptor signaling pathway
      negative regulation of protein complex assembly
      negative regulation of ossification
      ossification
      heparin binding
    • UniProt
    • Entrez Gene
    • Acknowledgements
      This antibody was developed by Bio-Rad, Zeppelinstr. 4, 82178 Puchheim, Germany, a wholly owned subsidiary of Bio-Rad Laboratories Inc.
    • Licensed Use
      For in vitro research purposes only, unless otherwise specified in writing by Bio-Rad.
    • Regulatory
      For research purposes only
    • This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visit the antibody protocols page.

    • Application NameYesNoMin DilutionMax Dilution
      Functional Assays
      Immunohistology - Frozen

    • Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own system using appropriate negative/positive controls.
    • Technical Advice
      Recommended protocols and further information about HuCAL recombinant antibody technology can be found in the HuCAL Antibodies Technical Manual
    • Recommended Protocol
    • ELISA
    • Immunohistology
    • Histology Positive Control Tissue
    • Immunofluorescence
    • Western Blotting
    • Instructions For Use

    Additional Sclerostin Antibody Formats

    Formats Clone Applications Sizes available
    Sclerostin Antibody : Preservative Free AbD09097_h/mIgG2a C, FN 0.1 mg
    • Copyright © 2016 Bio-Rad

    Recommended Secondary Antibody

      Recommended Negative Isotype Control

        Useful Reagents

          Recommended Positive Controls

            Histology Controls

              • Application NameReference Images
                Functional Assays

              References

              Further Reading

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