Anti-Calcium Pump pan PMCA ATPase antibody [5F10] (ab2825)

Overview

  • Product name
    Anti-Calcium Pump pan PMCA ATPase antibody [5F10]
  • Description
    Mouse monoclonal [5F10] to Calcium Pump pan PMCA ATPase
  • Tested applications
    Suitable for: ICC/IF, WB, ICC, IHC-P, Flow Cyt, Inhibition Assay, ELISA, IHC-Fr, IPmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Sheep, Rabbit, Chicken, Hamster, Cow, Cat, Dog, Human, Amphibian, Syrian hamster
    Predicted to work with: Non human primates
  • Immunogen

    Full length native protein (purified) corresponding to Human Calcium Pump pan PMCA ATPase. Purified human erythrocyte calcium ATPase.

  • Epitope
    This antibody recognizes an epitope between amino acids 724-783 of the human erythrocyte calcium pump.

Properties

Applications

Our Abpromise guarantee covers the use of ab2825 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 Use at an assay dependent concentration. PubMed: 17478566
WB 1/1000. This antibody recognizes an ~140 kDa protein representing PMCA ATPase and bands at 95kDa and 180 kDa which probably represent products of aggregation and/or natural proteolytic products of the pump from rat liver membrane preparations.
ICC Use at an assay dependent concentration.
IHC-P 1/500. Staining of PMCA ATPase yields a pattern consistent with that seen in the literature and depends on the tissue being studied and the localization of the isoforms present.
Flow Cyt 1/20 - 1/100.
Inhibition Assay Use at an assay dependent concentration.
ELISA Use at an assay dependent concentration.
IHC-Fr 1/500.
IP Use at an assay dependent concentration.

Target

  • Function
    This magnesium-dependent enzyme catalyzes the hydrolysis of ATP coupled with the transport of calcium out of the cell.
  • Tissue specificity
    Isoform B is ubiquitously expressed. Isoform C is found in brain cortex, skeletal muscle and heart muscle. Isoform D has only been found in fetal skeletal muscle. Isoform K has been found in small intestine and liver.
  • Sequence similarities
    Belongs to the cation transport ATPase (P-type) (TC 3.A.3) family. Type IIB subfamily.
  • Domain
    The calmodulin-binding subdomain B is different in the different splice variants and shows pH dependent calmodulin binding properties in isoforms A, C, D and E.
  • Cellular localization
    Cell membrane.
  • Information by UniProt
  • Database links
  • Alternative names
    • AT2B1_HUMAN antibody
    • ATP2B1 antibody
    • ATP2B2 antibody
    • ATP2B3 antibody
    • ATP2B4 antibody
    • ATPase Ca++ transporting plasma membrane 1 antibody
    • ATPase Ca++ transporting plasma membrane 2 antibody
    • ATPase Ca++ transporting plasma membrane 3 antibody
    • ATPase Ca++ transporting plasma membrane 4 antibody
    • Plasma membrane calcium ATPase isoform 1 antibody
    • Plasma membrane calcium pump isoform 1 antibody
    • Plasma membrane calcium transporting ATPase 1 antibody
    • Plasma membrane calcium transporting ATPase 2 antibody
    • Plasma membrane calcium transporting ATPase 3 antibody
    • Plasma membrane calcium transporting ATPase 4 antibody
    • Plasma membrane calcium-transporting ATPase 1 antibody
    • PMCA1 antibody
    • PMCA2 antibody
    • PMCA3 antibody
    • PMCA4 antibody
    see all

Images

  • All lanes : Anti-Calcium Pump pan PMCA ATPase antibody [5F10] (ab2825)

    Lane 1 : U251
    Lane 2 : Human brain
    Lane 3 : C2C12

    Lysates/proteins at 25 µg per lane.

    Secondary
    HRP-conjugated secondary antibody
  • Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human colon carcinoma tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:100 with a mouse monoclonal antibody recognizing PMCA ATPase ab2825 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

  • Immunocytochemistry/Immunofluorescence analysis of Calcium Pump pan PMCA ATPase shows staining in U251 cells. Calcium Pump pan PMCA ATPase staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or ab2825 (1:200) overnight at 4°C, washed with PBS and incubated with a DyLight-488 conjugated goat anti-mouse secondary antibody. Images were taken at 60X magnification.

  • Overlay histogram showing Jurkat cells stained with ab2825 (red line). The cells were fixed with 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 (ab2825, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG2a [ICIGG2A] (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in Jurkat cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween used with ab2825 (1/20 dilution) under the same conditions.

  • ab2825 Immunoprecipitate Calcium Pump pan PMCA ATPase in human whole cell lysate. 1,000,000 cells were lysed and incubated with primary antibody at 4µg/mg lysate and Protein A matrix for 20 hours at 4°C. For western blotting an undiluted Abcam`s ab1162, Rabbit polyclonal to DDDDK tag was used.

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  • Immunocytochemistry/Immunofluorescence analysis of Calcium Pump pan PMCA ATPase shows staining in C6 cells. Calcium Pump pan PMCA ATPase staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or ab2825 (1:200) overnight at 4°C, washed with PBS and incubated with a DyLight-488 conjugated goat anti-mouse secondary antibody. Images were taken at 60X magnification.

  • Immunocytochemistry/Immunofluorescence analysis of Calcium Pump pan PMCA ATPase shows staining in HeLa cells. Calcium Pump pan PMCA ATPase staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or ab2825 (1:100) overnight at 4°C, washed with PBS and incubated with a DyLight-488 conjugated goat anti-mouse secondary antibody. Images were taken at 60X magnification.

  • ab2825 at a 1/200 dilution detecting Calcium Pump pan PMCA ATPase in human monocytes by Flow Cytometry. The secondary used was an Alexa-Fluor 488 conjugated goat anti-mouse IgG (H+L) used at a 1/500 dilution.

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  • ab2825 staining Calcium Pump pan PMCA ATPase in human monocytes by Immunocytochemistry/ Immunofluorescence.
    Cells were fixed, permeabilized, blocked with 2% BSA for 30 minutes at 25°C and then incubated with ab2825 at a 1/250 dilution for 1 hour at 25°C. The secondary used was an Alexa-Fluor 488 conjugated goat anti-mouse IgG (H+L) used at a 1/1000 dilution.

    See Abreview

  • ab2825 staining Calcium Pump pan PMCA ATPase (green) in Mouse RAW 264.7 cells by ICC/IF (Immunocytochemistry/ Immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized in 0.1% Triton X-100 in 2% BSA for 15 minutes and blocked with 2% BSA for 1 hour at 22°C. Samples were incubated with primary antibody (1/200 in PBS + 2% BSA) for 18 hours at 4°C. An Alexa Fluor®488-conjugated Chicken anti-rabbit IgG polyclonal (H&L) (1/750) was used as the secondary antibody.
    WGA (red) = Wheat Germ Agglutinin

    See Abreview

  • Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human brain tissue tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:200 with a mouse monoclonal antibody recognizing PMCA ATPase ab2825 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

  • Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human tonsil tissue tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:200 with a mouse monoclonal antibody recognizing PMCA ATPase ab2825 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

References

This product has been referenced in:
  • Chang HY  et al. Selective serotonin reuptake inhibitor, fluoxetine, impairs E-cadherin-mediated cell adhesion and alters calcium homeostasis in pancreatic beta cells. Sci Rep 7:3515 (2017). Mouse . Read more (PubMed: 28615694) »
  • Chen J  et al. Besides an ITIM/SHP-1-dependent pathway, CD22 collaborates with Grb2 and plasma membrane calcium-ATPase in an ITIM/SHP-1-independent pathway of attenuation of Ca2+i signal in B cells. Oncotarget 7:56129-56146 (2016). Read more (PubMed: 27276708) »

See all 17 Publications for this product

Customer reviews and Q&As

The page 132 in the following book can provide further insight in ot the request you have made; https://books.google.co.uk/books?id=zXXSBwAAQBAJ&pg=PA133&lpg=PA133&dq=plasma+membrane+and+mitochondrial+membrane+marker&source=bl&ot...

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This antibody recognizes an epitope between amino acids 724-783 of the human erythrocyte calcium pump. These amino acids compose the highly conserved hinge region on th...

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (RAW 264.7)
Specification
RAW 264.7
Fixative
Paraformaldehyde
Permeabilization
Yes - 0.1% Triton-X100 in 2% BSA for 15min
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 22°C
Username

Dr. Mahesh Shivananjappa

Verified customer

Submitted May 24 2012

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Cell lysate - whole cell (Mouse retina)
Loading amount
25 µg
Specification
Mouse retina
Gel Running Conditions
Reduced Denaturing (8%)
Blocking step
Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Herr Dr. Vladimir Milenkovic

Verified customer

Submitted May 01 2012

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry
Sample
Human Cell (Monocytes from Apheresis)
Specification
Monocytes from Apheresis
Fixative
BD Cytofix/Cytoperm 20 min
Permeabilization
Yes - BD Cytofix/Cytoperm 20 min
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 2% · Temperature: 25°C
Username

Dr. Mahesh Shivananjappa

Verified customer

Submitted Feb 06 2012

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Flow Cytometry
Sample
Human Cell (Monocytes from Apheresis)
Specification
Monocytes from Apheresis
Fixation
BD Cytofix/Cytoperm 20 min
Permeabilization
Yes - BD Cytofix/Cytoperm 20 min
Gating Strategy
Monocytes
Username

Dr. Mahesh Shivananjappa

Verified customer

Submitted Dec 07 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Cell lysate - whole cell (EL4 cell line)
Loading amount
20000 cells
Specification
EL4 cell line
Treatment
cells expressing recombitant viral protein
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 minute(s) · Concentration: 0.1% · Temperature: 20°C
Username

Abcam user community

Verified customer

Submitted Jul 07 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Tissue lysate - other (Synaptosomes)
Loading amount
20 µg
Specification
Synaptosomes
Gel Running Conditions
Reduced Denaturing
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted Aug 13 2009

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (huh-7)
Loading amount
20000 cells
Specification
huh-7
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 minute(s) · Concentration: 0.1% · Temperature: 20°C
Username

Abcam user community

Verified customer

Submitted Jul 29 2009

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunoprecipitation
Sample
Human Cell lysate - whole cell (vero)
Total protein in input
1e+006 cells
Specification
vero
Treatment
cells expressing recombitant protein tagged with FLAG
Immuno-precipitation step
Protein A
Username

Abcam user community

Verified customer

Submitted Jul 29 2009

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