Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-Calnexin antibody [EPR3632] (ab92573)

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Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
  • Immunocytochemistry/ Immunofluorescence - Anti-Calnexin antibody [EPR3632] (ab92573)
  • Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
  • Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calnexin antibody [EPR3632] (ab92573)
  • Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
  • Anti-Calnexin antibody [EPR3632] (ab92573)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR3632] to Calnexin
  • Suitable for: ICC/IF, WB, IP, IHC-P
  • Knockout validated
  • Reacts with: Mouse, Human

Overview

  • Product name

    Anti-Calnexin antibody [EPR3632]
    See all Calnexin primary antibodies

  • Description

    Rabbit monoclonal [EPR3632] to Calnexin

  • Host species

    Rabbit

  • Specificity

    Recognizes ER membrane, mitochondria and cis-Golgi

  • Tested applications

    Suitable for: ICC/IF, WB, IP, IHC-Pmore details
    Unsuitable for: Flow Cyt

  • Species reactivity

    Reacts with: Mouse, Human

  • Immunogen

    Synthetic peptide within Human Calnexin aa 1-100. The exact sequence is proprietary.
    Database link: P27824

  • Positive control

    • WB: HeLa, A431, SH-SY5Y, HEK-293T, MCF7, U-2 OS and HepG2 whole cell lysate (ab7900). IHC-P: Human tonsil tissue. ICC/IF: Wild-type HAP1 cells.

  • General notes

    Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.

    References regarding specificity:

    Horner SM  et al. Mitochondrial-associated endoplasmic reticulum membranes (MAM) form innate immune synapses and are targeted by hepatitis C virus. Proc Natl Acad Sci U S A 108:14590-5 (2011). PubMed: 21844353

    Myhill N  et al. The subcellular distribution of calnexin is mediated by PACS-2. Mol Biol Cell 19:2777-88 (2008). PubMed: 18417615

    Yoshimura SI  et al. Direct targeting of cis-Golgi matrix proteins to the Golgi apparatus. J Cell Sci 114:4105-15 (2001). PubMed: 11739642

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

Properties

  • Form

    Liquid

  • Storage instructions

    Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid repeated freeze / thaw cycles.

  • Storage buffer

    pH: 7.20
    Preservative: 0.05% Sodium azide
    Constituents: 0.1% BSA, 40% Glycerol (glycerin, glycerine), 9.85% Tris glycine, 50% Tissue culture supernatant
  • Concentration information loading...

  • Purity

    Tissue culture supernatant

  • Clonality

    Monoclonal

  • Clone number

    EPR3632

  • Isotype

    IgG

  • Research areas

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab92573 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
ICC/IF
1/1000.
WB
1/20000 - 1/100000. Predicted molecular weight: 90 kDa.
IP
1/50.
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Notes
ICC/IF
1/1000.
WB
1/20000 - 1/100000. Predicted molecular weight: 90 kDa.
IP
1/50.
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
  • Application notes
    Is unsuitable for Flow Cyt.

    • Target

    • Function

      Calcium-binding protein that interacts with newly synthesized glycoproteins in the endoplasmic reticulum. It may act in assisting protein assembly and/or in the retention within the ER of unassembled protein subunits. It seems to play a major role in the quality control apparatus of the ER by the retention of incorrectly folded proteins.

    • Sequence similarities

      Belongs to the calreticulin family.

    • Cellular localization

      Endoplasmic reticulum membrane. Melanosome. Identified by mass spectrometry in melanosome fractions from stage I to stage IV.
    • Information by UniProt

    • Database links

      • Alternative names

        • Calnexin antibody
        • CALX_HUMAN antibody
        • CANX antibody
        • CNX antibody
        • FLJ26570 antibody
        • Histocompatibility complex class I antigen binding protein p88 antibody
        • IP90 antibody
        • Major histocompatibility complex class I antigen-binding protein p88 antibody
        • p90 antibody
        see all

      Images

      • Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
        Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
        All lanes : Anti-Calnexin antibody [EPR3632] (ab92573) at 1/20000 dilution

        Lane 1 : Wild-type HEK-293T cell lysate
        Lane 2 : CANX knockout HEK-293T cell lysate
        Lane 3 : U-2 OS cell lysate
        Lane 4 : MCF7 cell lysate

        Lysates/proteins at 20 µg per lane.

        Performed under reducing conditions.

        Predicted band size: 90 kDa
        Observed band size: 80 kDa why is the actual band size different from the predicted?



        Lanes 1 - 4: Merged signal (red and green). Green - ab92573 observed at 80 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.

        ab92573 was shown to react with Calnexin in wild-type HEK-293T cells in Western blot with loss of signal observed in CANX knockout cell line ab255368 (CANX knockout cell lysate ab263805). Wild-type HEK-293T and CANX knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab92573 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4 °C at a 1 in 20000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.

      • Immunocytochemistry/ Immunofluorescence - Anti-Calnexin antibody [EPR3632] (ab92573)
        Immunocytochemistry/ Immunofluorescence - Anti-Calnexin antibody [EPR3632] (ab92573)

        ab92573 staining Calnexin in wild-type HAP1 cells (top panel) and CANX knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab92573 at 1/1000 dilution and ab195889 at 1/250 dilution (shown in pseudocolour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

        Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

      • Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
        Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
        All lanes : Anti-Calnexin antibody [EPR3632] (ab92573) at 1/20000 dilution

        Lane 1 : Wild-type HAP1 cell lysate
        Lane 2 : Calnexin knockout HAP1 cell lysate
        Lane 3 : THP-1 cell lysate
        Lane 4 : RAW 264.7 cell lysate

        Lysates/proteins at 20 µg per lane.

        Predicted band size: 90 kDa



        Lanes 1 - 4: Merged signal (red and green). Green - ab92573 observed at 80 kDa. Red - loading control, ab8245, observed at 37 kDa.

        ab92573 was shown to specifically react with Calnexin when Calnexin knockout samples were used. Wild-type and Calnexin knockout samples were subjected to SDS-PAGE. ab92573 and ab8245 (loading control to GAPDH) were diluted at 1/20,000 and 1/10,000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776 secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

      • Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
        Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
        All lanes : Anti-Calnexin antibody [EPR3632] (ab92573) at 1/20000 dilution

        Lane 1 : HeLa cell lysate
        Lane 2 : A431 cell lysate
        Lane 3 : SH-SY5Y cell lysate
        Lane 4 : HepG2 cell lysates

        Lysates/proteins at 10 µg per lane.

        Secondary
        All lanes : standard HRP labelled goat anti-rabbit at 1/2000 dilution

        Predicted band size: 90 kDa

      • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calnexin antibody [EPR3632] (ab92573)
        Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calnexin antibody [EPR3632] (ab92573)

        Immunohistochemical analysis of paraffin embedded Human tonsil tissue using ab92573 at a 1/100 dilution.

        Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

      • Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
        Western blot - Anti-Calnexin antibody [EPR3632] (ab92573)
        All lanes : Anti-Calnexin antibody [EPR3632] (ab92573)

        Lane 1 : Wild-type HAP1 cell lysate
        Lane 2 : Calnexin knockout HAP1 cell lysate

        Lysates/proteins at 20 µg per lane.

        Predicted band size: 90 kDa



        Lanes 1 - 2: Merged signal (red and green). Green - ab92573 observed at 80 kDa. Red - loading control, ab8245, observed at 37 kDa.

        This western blot image is a comparison between ab92573 and a competitor's rabbit polyclonal antibody.

      • Anti-Calnexin antibody [EPR3632] (ab92573)
        Anti-Calnexin antibody [EPR3632] (ab92573)

      References (9)

      Publishing research using ab92573? Please let us know so that we can cite the reference in this datasheet.

      ab92573 has been referenced in 9 publications.

      • Xie M  et al. Exosomal circSHKBP1 promotes gastric cancer progression via regulating the miR-582-3p/HUR/VEGF axis and suppressing HSP90 degradation. Mol Cancer 19:112 (2020). PubMed: 32600329
      • Yan L & Wu X Exosomes produced from 3D cultures of umbilical cord mesenchymal stem cells in a hollow-fiber bioreactor show improved osteochondral regeneration activity. Cell Biol Toxicol 36:165-178 (2020). PubMed: 31820164
      • Zhu J  et al. Exosomes from nicotine-stimulated macrophages accelerate atherosclerosis through miR-21-3p/PTEN-mediated VSMC migration and proliferation. Theranostics 9:6901-6919 (2019). PubMed: 31660076
      • Anczurowski M  et al. Chaperones of the class I peptide-loading complex facilitate the constitutive presentation of endogenous antigens on HLA-DP84GGPM87. J Autoimmun 102:114-125 (2019). PubMed: 31078377
      • Moonschi FH  et al. Mammalian Cell-derived Vesicles for the Isolation of Organelle Specific Transmembrane Proteins to Conduct Single Molecule Studies. Bio Protoc 8:N/A (2018). PubMed: 30406159
      • Baba K  et al. Asiatic Acid, Corosolic Acid, and Maslinic Acid Interfere with Intracellular Trafficking and N-Linked Glycosylation of Intercellular Adhesion Molecule-1. Biol Pharm Bull 41:1757-1768 (2018). PubMed: 30504678
      • Fox-Loe AM  et al. Organelle-specific single-molecule imaging of a4ß2 nicotinic receptors reveals the effect of nicotine on receptor assembly and cell-surface trafficking. J Biol Chem 292:21159-21169 (2017). PubMed: 29074617
      • Shwetha S  et al. HuR Displaces Polypyrimidine Tract Binding Protein To Facilitate La Binding to the 3' Untranslated Region and Enhances Hepatitis C Virus Replication. J Virol 89:11356-71 (2015). WB ; Human . PubMed: 26339049
      • Mitsuda S  et al. Ursolic acid, a natural pentacyclic triterpenoid, inhibits intracellular trafficking of proteins and induces accumulation of intercellular adhesion molecule-1 linked to high-mannose-type glycans in the endoplasmic reticulum. FEBS Open Bio 4:229-39 (2014). ICC/IF ; Human . PubMed: 24649404

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      Western blot abreview for Anti-Calnexin antibody [EPR3632]

       Inconclusive
      Abreviews
      abreview image
      Application
      Western blot
      Sample
      Mouse Cell lysate - whole cell (catecholaminergic neuronal tumor)
      Gel Running Conditions
      Reduced Denaturing (NuPAGE 4-12% Bis-Tris, 15 wells)
      Loading amount
      10 µg
      Treatment
      Chronically infected with RML prion strain
      Specification
      catecholaminergic neuronal tumor
      Blocking step
      Milk as blocking agent for 1 hour(s) and 30 minute(s) · Concentration: 5% · Temperature: 4°C
      Read More

      Dr. Manjeet Kumar

      Verified customer

      Submitted Mar 15 2019

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