Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-Calnexin antibody [EPR3633(2)] - BSA and Azide free (ab225542)

Overview

  • Product name

    Anti-Calnexin antibody [EPR3633(2)] - BSA and Azide free
    See all Calnexin primary antibodies
  • Description

    Rabbit monoclonal [EPR3633(2)] to Calnexin - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, IHC-P, Flow Cytmore details
  • Species reactivity

    Reacts with: Rat, Human
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) corresponding to Human Calnexin aa 550-650 (C terminal).

  • Positive control

    • HepG2, A431, SH-SY5Y and HeLa cell lysates, Human kidney tissue
  • General notes

    ab225542 is the carrier-free version of ab133615 This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    Use our conjugation kits  for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    Ab225542 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

    Mouse: 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

    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

Applications

Our Abpromise guarantee covers the use of ab225542 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
WB Use at an assay dependent concentration. Detects a band of approximately 90 kDa (predicted molecular weight: 68 kDa).
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

Flow Cyt Use at an assay dependent concentration.

ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

 

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

  • This WB data was generated using the same anti-Calnexin antibody clone [EPR3633(2)] in a different buffer formulation (cat# ab133615).

    Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: Calnexin knockout HAP1 cell lysate (20 µg) 
    Lane 3: THP1 cell lysate (20 µg)
    Lane 4: Raw264.7 cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab133615 observed at 80 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab133615 was shown to recognize Calnexin when Calnexin knockout samples were used, along with additional cross-reactive bands. Wild-type and Calnexin knockout samples were subjected to SDS-PAGE. ab133615 and ab8245 (loading control to GAPDH) were diluted 1/1000 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 h at room temperature before imaging.

  • Overlay histogram showing HeLa cells fixed in 80% methanol and stained with purified ab133615 at a dilution of 1 in 360 (red line). The secondary antibody used was FITC goat anti-rabbit at a dilution of 1 in 150. Rabbit monoclonal IgG was used as an isotype control (black line) and cells without incubation with antibody were used as a negative control (blue line).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab133615).

  • Immunohistochemical staining of paraffin embedded rat cardiac muscle with purified ab133615 at a working dilution of 1 in 4000. The secondary antibody used is ab97051, a HRP goat anti-rabbit (H+L), at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab133615).

  • Immunohistochemical staining of paraffin embedded human pancreas with purified ab133615 at a working dilution of 1 in 4000. The secondary antibody used is ab97051, a HRP goat anti-rabbit (H+L), at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab133615).

  • Overlay histogram showing HeLa cells stained with unpurified ab133615 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (unpurified ab133615, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in HeLa cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab133615).

  • Immunohistochemical analysis of paraffin-embedded human kidney tissue labelled with unpurified ab133615 at 1/100 dilution.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab133615).

    Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

References

ab225542 has not yet been referenced specifically in any publications.

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