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ab23379 |
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ab23379 |
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Read our guarantee »Products:Tags & Cell Markers >> Subcellular Markers >> Organelles >> ER
Anti-Calnexin - ER membrane marker antibody
See all Calnexin - ER membrane marker products (12) ...
Rabbit polyclonal to Calnexin - ER membrane marker
WB, ICC/IF, IHC-P, IPmore details
Reacts with
Mouse, Rat, Human, Marmoset (common)
Predicted to work with
Dog
Synthetic peptide conjugated to KLH derived from within residues 550 to the C-terminus of Human Calnexin.
(Peptide available as ab23379.)
ab22595 gave a positive result in the following Human Whole Cell Lysates: Hela, MCF-7 Mouse Whole Cell Lystates: NIH 3T3, MEF1 Mouse Tissue Lysates: Brain, Liver, Heart, Kidney, Pancreas, Testis, Skeletal muscle, Spinal cord, Ovary Rat Whole Cell Lysates: PC12, Rat Tissue Lysates: Brain, Liver, Heart, Kidney
Liquid
Store at +4°C short term (1-2 weeks). Aliquot and store at -20°C or -80°C. Store in the dark. Avoid repeated freeze / thaw cycles.
Preservative: 0.02% Sodium Azide
Constituents: 1% BSA, PBS, pH 7.4
Concentration information loading...
Immunogen affinity purified
Polyclonal
IgG
Neuroscience >> Sensory System >> Visual system
Tags & Cell Markers >> Subcellular Markers >> Organelles >> ER
Our Abpromise guarantee covers the use of ab22595 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
WB: Use a concentration of 1 µg/mlDetects a band of approximately 75 kDa (predicted molecular weight: 90 kDa).Can be blocked with Calnexin peptide (ab23379).
ICC/IF: Use a concentration of 5 µg/ml
IHC-P: Use at an assay dependent concentration.
IP: Use at an assay dependent concentration.
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.
Belongs to the calreticulin family.
Endoplasmic reticulum membrane. Melanosome. Identified by mass spectrometry in melanosome fractions from stage I to stage IV.
Target information above from: UniProt accessionP27824
The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010).
Immunocytochemistry/ Immunofluorescence - Calnexin antibody (ab22595)

ICC/IF image of ab22595 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab22595, 5µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).
Western blot - Calnexin antibody (ab22595)

All lanes : Anti-Calnexin - ER membrane marker antibody (ab22595) at 1 µg/ml
Lane 1 : HeLa (Human epithelial carcinoma cell line)
Lane 2 : U2OS Whole Cell Lysate
Lane 3 : MCF-7 (Human breast adenocarcinoma cell line) Whole Cell Lysate
Lane 4 : HeLa (Human epithelial carcinoma cell line) with Calnexin peptide (ab23379) at 1 µg/ml
Lane 5 : U2OS Whole Cell Lysate with Calnexin peptide (ab23379) at 1 µg/ml
Lane 6 : MCF-7 (Human breast adenocarcinoma cell line) Whole Cell Lysate with Calnexin peptide (ab23379) at 1 µg/ml
Lysates/proteins at 20 µg per lane.
Secondary
Goat polyclonal to Rabbit IgG (Alexa Fluor® 680) at 1/10000 dilution
Performed under reducing conditions.
Predicted band size : 90 kDa
Observed band size : 75 kDa (why is the actual band size different from the predicted?)
Recent batches of ab22595 (AP217379 and AP151845) detect a band of ~ 75 kDa in Hela, U2OS and MCF-7 lysates. This band is completely blocked by the immunizing peptide so we believe this represents Calnexin. Moreoever, a band of the same size is detected by other Calnexin antibodies tested.
Western blot - Calnexin - ER membrane marker antibody (ab22595)

All lanes : Anti-Calnexin - ER membrane marker antibody (ab22595) at 1/250 dilution
Lane 1 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate (ab7179)
Lane 2 : MEF1 (Mouse embryonic fibroblast cell line) Whole Cell Lysate (ab46770)
Lane 3 : Brain (Mouse) Tissue Lysate (ab27253)
Lane 4 : Liver (Mouse) Tissue Lysate (ab7935)
Lane 5 : Heart (Mouse) Tissue Lysate (ab27255)
Lane 6 : Kidney (Mouse) Tissue Lysate (ab27254)
Lane 7 : Pancreas (Mouse) Tissue Lysate (ab29363)
Lane 8 : Testis (Mouse) Tissue Lysate - normal tissue (ab4027)
Lane 9 : Skeletal Muscle (Mouse) Tissue Lysate (ab29711)
Lane 10 : Spinal Cord (Mouse) Tissue Lysate (ab50253)
Lane 11 : Ovary (Mouse) Tissue Lysate (ab35808)
Lane 12 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate (ab50957)
Lane 13 : Brain (Rat) Tissue Lysate (ab7942)
Lane 14 : Liver (Rat) Tissue Lysate (ab27256)
Lane 15 : Heart (Rat) Tissue Lysate (ab7940)
Lane 16 : Kidney (Rat) Whole Cell Lysate - normal tissue (ab29480)
Lysates/proteins at 10 µg per lane.
Secondary
IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution
Performed under reducing conditions.
Predicted band size : 90 kDa
Observed band size : 80 kDa (why is the actual band size different from the predicted?)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Calnexin - ER membrane marker antibody (ab22595)

Kidney cortex using ab22595 shows clear cytoplasmic staining patterns. The visceral cells of the Glomerular tuft ( podocytes ) are strongly stained (indicated by red arrowheads). Distal convoluted tubular cells are generally moderately positive (with exceptions that are strongly positive). However, most of the cells that line the Proximal Convoluted Tubules (indicated by green arrowheads) are strongly positive.
Carl Hobbs, King`s College London, United Kingdom
Immunocytochemistry/ Immunofluorescence - Calnexin - ER membrane marker antibody (ab22595)

ICC/IF image of ab22595 stained HeLa cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab22595, 5µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-rabbit IgG - H&L, pre-adsorbed (ab96899) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
Immunoprecipitation - Anti-Calnexin - ER membrane marker antibody (ab22595)

Calnexin - ER membrane marker was immunoprecipitated using 0.5mg Hela whole cell extract, 5ug of Rabbit polyclonal to Calnexin - ER membrane markerand 50µl of protein G magnetic beads (lane 1). The antibody was incubated with the Protein G beads for 10min under agitation. No antibody was added to the control (lane 2). Hela whole cell extractdiluted in RIPA buffer was added to each sample and incubated for 10min under agitation. Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab22595. Secondary: Mouse monoclonal [SB62a] to rabbit IgG light chain specific (HRP) (ab99697) at 1/5000 dilution. Band: 80kDa: Calnexin - ER membrane marker.
This product has been referenced in:
See all 7 publications for this product
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ICC/IF image of ab22595 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab22595, 5µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).

All lanes : Anti-Calnexin - ER membrane marker antibody (ab22595) at 1 µg/ml
Lane 1 : HeLa (Human epithelial carcinoma cell line)
Lane 2 : U2OS Whole Cell Lysate
Lane 3 : MCF-7 (Human breast adenocarcinoma cell line) Whole Cell Lysate
Lane 4 : HeLa (Human epithelial carcinoma cell line) with Calnexin peptide (ab23379) at 1 µg/ml
Lane 5 : U2OS Whole Cell Lysate with Calnexin peptide (ab23379) at 1 µg/ml
Lane 6 : MCF-7 (Human breast adenocarcinoma cell line) Whole Cell Lysate with Calnexin peptide (ab23379) at 1 µg/ml
Lysates/proteins at 20 µg per lane.
Secondary
Goat polyclonal to Rabbit IgG (Alexa Fluor® 680) at 1/10000 dilution
Performed under reducing conditions.
Predicted band size : 90 kDa
Observed band size : 75 kDa (why is the actual band size different from the predicted?)
Recent batches of ab22595 (AP217379 and AP151845) detect a band of ~ 75 kDa in Hela, U2OS and MCF-7 lysates. This band is completely blocked by the immunizing peptide so we believe this represents Calnexin. Moreoever, a band of the same size is detected by other Calnexin antibodies tested.

All lanes : Anti-Calnexin - ER membrane marker antibody (ab22595) at 1/250 dilution
Lane 1 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate (ab7179)
Lane 2 : MEF1 (Mouse embryonic fibroblast cell line) Whole Cell Lysate (ab46770)
Lane 3 : Brain (Mouse) Tissue Lysate (ab27253)
Lane 4 : Liver (Mouse) Tissue Lysate (ab7935)
Lane 5 : Heart (Mouse) Tissue Lysate (ab27255)
Lane 6 : Kidney (Mouse) Tissue Lysate (ab27254)
Lane 7 : Pancreas (Mouse) Tissue Lysate (ab29363)
Lane 8 : Testis (Mouse) Tissue Lysate - normal tissue (ab4027)
Lane 9 : Skeletal Muscle (Mouse) Tissue Lysate (ab29711)
Lane 10 : Spinal Cord (Mouse) Tissue Lysate (ab50253)
Lane 11 : Ovary (Mouse) Tissue Lysate (ab35808)
Lane 12 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate (ab50957)
Lane 13 : Brain (Rat) Tissue Lysate (ab7942)
Lane 14 : Liver (Rat) Tissue Lysate (ab27256)
Lane 15 : Heart (Rat) Tissue Lysate (ab7940)
Lane 16 : Kidney (Rat) Whole Cell Lysate - normal tissue (ab29480)
Lysates/proteins at 10 µg per lane.
Secondary
IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution
Performed under reducing conditions.
Predicted band size : 90 kDa
Observed band size : 80 kDa (why is the actual band size different from the predicted?)

Kidney cortex using ab22595 shows clear cytoplasmic staining patterns. The visceral cells of the Glomerular tuft ( podocytes ) are strongly stained (indicated by red arrowheads). Distal convoluted tubular cells are generally moderately positive (with exceptions that are strongly positive). However, most of the cells that line the Proximal Convoluted Tubules (indicated by green arrowheads) are strongly positive.
Carl Hobbs, King`s College London, United Kingdom

ICC/IF image of ab22595 stained HeLa cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab22595, 5µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-rabbit IgG - H&L, pre-adsorbed (ab96899) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

Calnexin - ER membrane marker was immunoprecipitated using 0.5mg Hela whole cell extract, 5ug of Rabbit polyclonal to Calnexin - ER membrane markerand 50µl of protein G magnetic beads (lane 1). The antibody was incubated with the Protein G beads for 10min under agitation. No antibody was added to the control (lane 2). Hela whole cell extractdiluted in RIPA buffer was added to each sample and incubated for 10min under agitation. Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab22595. Secondary: Mouse monoclonal [SB62a] to rabbit IgG light chain specific (HRP) (ab99697) at 1/5000 dilution. Band: 80kDa: Calnexin - ER membrane marker.
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