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JAG1 Antikörper (Jagged 1) Internal Region Primary Antibody

JAG1 Reaktivität: Human, Maus FM, IHC, ELISA, WB Wirt: Kaninchen Polyclonal unconjugated
Pubmed (2 references) Independent Validation (1 Validierung)
Produktnummer ABIN129524
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  • Target Alle JAG1 Antikörper anzeigen
    JAG1
    Bindungsspezifität
    • 20
    • 9
    • 8
    • 5
    • 5
    • 4
    • 3
    • 3
    • 3
    • 3
    • 2
    • 2
    • 2
    • 2
    • 2
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    Internal Region
    Reaktivität
    • 90
    • 29
    • 25
    • 1
    Human, Maus
    Wirt
    • 67
    • 21
    • 7
    • 4
    Kaninchen
    Klonalität
    • 69
    • 30
    Polyklonal
    Konjugat
    • 47
    • 11
    • 9
    • 6
    • 5
    • 4
    • 3
    • 2
    • 2
    • 2
    • 2
    • 2
    • 2
    • 1
    • 1
    Dieser JAG1 Antikörper ist unkonjugiert
    Applikation
    • 61
    • 48
    • 34
    • 27
    • 9
    • 8
    • 8
    • 6
    • 3
    • 2
    • 2
    • 2
    • 1
    Fluorescence Microscopy (FM), Immunohistochemistry (IHC), ELISA, Western Blotting (WB)
    Spezifität
    This protein A purified antibody is directed against human Jagged-1 protein. The product was purified from mono-specific antiserum by affinity chromatography. A BLAST analysis was used to suggest cross reactivity with Jagged-1 protein from human, chimpanzee, rat and mouse based on 100% homology with the immunizing sequence. Partial reactivity is expected against dog (81%) and Xenopus laevis (85%) based on partial sequence homologies as indicated. Reactivity against homologues from other sources is not known.
    Produktmerkmale
    Anti Jagged 1 Antibody recognizes the jagged 1 protein encoded by the JAG1 gene, that is the human homolog of the Drosophilia jagged protein. Human jagged 1 is the ligand for the receptor notch 1, the latter a human homolog of the Drosophilia jagged receptor notch. Mutations that alter the jagged 1 protein cause Alagille syndrome. Jagged 1 signaling through notch 1 has also been shown to play a role in hematopoies.
    Aufreinigung
    purified
    Sterilität
    Sterile filtered
    Immunogen
    This protein A purified antibody was prepared from whole rabbit serum produced by repeated immunizations with a synthetic peptide corresponding to amino acids 110-125 of human Jagged-1protein.
    Immunogen Type: Peptide
    Isotyp
    IgG
  • Applikationshinweise
    This protein A purified antibody has been tested for use in ELISA, immunohistochemistry, immunofluorescence microscopy and western blot. Specific conditions for reactivity should be optimized by the end user. Expect a band approximately 150 kDa in size corresponding to Jagged-1 in mouse liver whole cell lysates, and a 75 kDa band in human brain and kidney lysates.
    Kommentare

    Gene Name: JAG1

    Beschränkungen
    Nur für Forschungszwecke einsetzbar
  • Validierung #103702 (Western Blotting)
    'Independent Validation' Siegel
    by
    AG Pancreatic Development and Stem cell differentiation, Universitätsklinikum Ulm
    No.
    #103702
    Datum
    16.06.2019
    Antigen
    JAG1
    Chargennummer
    28285
    Validierte Anwendung
    Western Blotting
    Positivkontrolle

    human pluripotent stem cells (day 0)

    pancreatic progenitor cells (day 14)

    Negativkontrolle

    definite endoderm cells (day 4)

    pancreatic endoderm cells (day 10)

    Bewertung

    ABIN1043767 reveals a protein band of the expected MW and some weaker extraneous bands in cell lysates of human pluripotent stem cells and pancreatic progenitor cells.

    'Independent Validation' Siegel
    Validierungsbilder
    Protokoll
    Primärantikörper
    ABIN1043767
    Sekundärantikörper
    donkey anti-rabbit HRP-conjugated antibody (GE Healthcare, NA9310V)
    Full Protocol
    • Grow HUES8 in mTeSR1 (STEMCELL Technologies, 85850) at 37°C and 5% O2, 5% CO2 in 2ml in a 6-well plate to 90% confluency.
    • Harvest cells using TrypLE (ThermoFisher Scientific, 12604013) following the manufacturer´s instructions.
    • Lyse 2x106 cells in 30µl per well cold RIPA buffer (50mM Tris-HCl pH 8.0, 150mM NaCl, 0.1% SDS, 0.5% deoxycholate, 1% TritonX 100 in ddH2O) supplemented with 1mM PMSF and 1x protease inhibitor (Roche, 11836170001) for 30min on ice with 3x vortexing in 1.5ml microcentrifuge tubes.
    • Centrifuge tubes at 10000xg for 8min at 4°C.
    • Transfer supernatant to a new 1.5ml microcentrifuge tube and store at -80°C.
    • Determine total protein content of the lysates using a Bradford assay (Bio-Rad, 500-0006).
    • Denature 50µg of total protein for 5min at 95°C in 30µl 1x Laemmli SDS sample buffer and subsequently separate them on a denaturing 7.5% polyacrylamide gel (7.5% Acrylamide, 0.375M Trís-HCl (pH8.8), 0.1% SDS, 0.1% APS, 0.1% TEMED) for about 90min at 120V.
    • Transfer proteins onto a PVDF membrane (Sigma Aldrich, IPVH00010) with transfer buffer (5.27g Tris, 2.93 g glycerine, 200 ml methanol, fill to 1l ddH2O) in a semidry western blotting system for 75min at 80mA/gel.
    • Check transfer of the separated proteins by Ponceau S staining.
    • Rinse membrane with water.
    • Wash membrane for 5min with TBST (TBS, 0.02% Tween20).
    • Block the membrane with 20ml blocking buffer (TBST, 5% milk) on a shaker for 1h at RT.
    • Rinse membrane 3x with TBST.
    • Wash membrane on a shaker 3x for 5min with TBST.
    • Shrink-wrap and incubate membrane with primary
      • rabbit anti-JAG1 antibody (antibodies-online, ABIN1043767, lot 28285) diluted 1:1000 and 1:10000 respectively in blocking buffer ON at 4°C or
      • mouse anti-beta actin antibody (Sigma Aldrich, A1978) diluted 1:2000 in blocking buffer at RT for 1h.
    • Wash membrane 3x for 5min with TBST.
    • Incubate membrane with secondary
      • donkey anti-rabbit HRP-conjugated antibody (GE Healthcare, NA9310V) diluted 1:5000 in TBST containing 1% milk for 1h at RT or
      • donkey anti-mouse HRP-conjugated antibody (GE Healthcare, NA9340V) diluted 1:5000 in TBST containing 1% milk for 1h at RT.
    • Wash membrane 3x for 5min with TBST.
    • Reveal protein bands using ECL solution (ThermoScientific, 34076) on a Fusion SL (Vilber) chemiluminescence system. Exposure times for the 1:1000 and 1:10000 dilutions of ABIN1043767 were 30sec and 5min respectively.
    Anmerkungen
    • The bands marked by the arrow in the western blot image using ABIN1043767 at a 1:1000 dilution seem to be specific, as JAG1 showed highest transcriptomic levels at pancreatic progenitor stage (lanes 7 and 8), followed by pluripotent stem cell state (lane 2). Although the immunoblot with ABIN1043767 results in some unspecific bands, the expected band height fits nicely (75kDa). Higher MW bands might potentially represent precursor proteins detectable in all samples but pancreatic progenitor stage samples. In principle specific bands are also detectable with ABIN1043767 at a 1:10000 dilution, although exposition time was very long.

    • ABIN1043767 was also tested in IHC on 4-5µm FFPE sections of primary human kidney cancer tissue. Epitope retrieval was carried out using Tris-EDTA buffer at pH9.0 (Zytomed, ZUC029-500), EDTA at pH8.0 (Leica, RE7116), or citrate buffer at pH6.1 (Agilent, S169984-2) for 20min in a decloaking chamber. Antigen retrieval at pH8.0 looks ok, assuming a broad expression pattern. However, detection would be rather expected in fewer tubular cells. No staining was observed in glomeruli. Cytoplasmic staining in tubuli is revealed at higher magnification. Note that antigen retrieval influences the staining pattern. All in all, staining needs to be validated for example in protein ablation assays.

  • Format
    Liquid
    Konzentration
    1.1 mg/mL
    Buffer
    0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2
    Vorsichtsmaßnahmen
    This product contains sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
    Lagerung
    4 °C/-20 °C
    Informationen zur Lagerung
    Store vial at 4 °C prior to restoration. For extended storage aliquot contents and freeze at -20 °C or below. Avoid cycles of freezing and thawing. Centrifuge product if not completely clear after standing at room temperature. This product is stable for several weeks at 4 °C as an undiluted liquid. Dilute only prior to immediate use. Expiration date is one (1) year from date of opening.
    Haltbarkeit
    12 months
  • Boareto, Jolly, Goldman, Pietilä, Mani, Sengupta, Ben-Jacob, Levine, Onuchic: "Notch-Jagged signalling can give rise to clusters of cells exhibiting a hybrid epithelial/mesenchymal phenotype." in: Journal of the Royal Society, Interface, Vol. 13, Issue 118, (2017) (PubMed).

    Yabe, Matsumoto, Tsurumoto, Shindo: "Immunohistological localization of Notch receptors and their ligands Delta and Jagged in synovial tissues of rheumatoid arthritis." in: Journal of orthopaedic science : official journal of the Japanese Orthopaedic Association, Vol. 10, Issue 6, pp. 589-94, (2005) (PubMed).

  • Target
    JAG1
    Andere Bezeichnung
    Jagged 1 (JAG1 Produkte)
    Synonyme
    AGS, AHD, AWS, CD339, HJ1, JAGL1, ABE2, Gsfabe2, Htu, Ozz, Ser-1, JAG1, jag3, cb243, jag1, sb:cb243, serrateC, jagged1, serrate, serrate-1, x-serrate-1, jagged 1, jagged 1b, jagged 1a, jagged 1 L homeolog, JAG1, Jag1, jag1, jag1b, jag1a, jag1.L
    Hintergrund
    Anti Jagged 1 Antibody recognizes the jagged 1 protein encoded by the JAG1 gene, that is the human homolog of the Drosophilia jagged protein. Human jagged 1 is the ligand for the receptor notch 1, the latter a human homolog of the Drosophilia jagged receptor notch. Mutations that alter the jagged 1 protein cause Alagille syndrome. Jagged 1 signaling through notch 1 has also been shown to play a role in hematopoies.
    Synonyms: Ser 1 antibody, AGS antibody, AHD antibody, AWS antibody, CD 339 antibody, CD339 antibody, CD339 antigen antibody, Headturner antibody, HJ1 antibody, Htu antibody
    Gen-ID
    182, 4557679
    UniProt
    P78504
    Pathways
    Notch Signalweg, Stem Cell Maintenance
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