Recombinant Anti-Calcineurin A antibody [EPR24997-22] KO Tested (ab282104)

Product name

Anti-Calcineurin A antibody [EPR24997-22]
See all Calcineurin A primary antibodies
Description

Rabbit monoclonal [EPR24997-22] to Calcineurin A
Host species

Rabbit
Tested applications

Suitable for: IP, ICC/IF, IHC-P, Flow Cyt (Intra), WBmore details
Species reactivity

Reacts with: Mouse, Rat, Human
Immunogen

Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: Wild-type HAP1, HeLa, SH-SY5Y, Mouse brain, Rat brain, Mouse lung, Rat lung, Human PPP3CA recombinant protein lysates. IHC-P: Human cerebrum, Human astrocytoma, Human colon carcinoma, Mouse cerebrum and Rat cerebrum tissues, Parental HAP1 cell pellet. ICC/IF: HeLa, NIH/3T3 cells. Flow Cyt: HeLa, NIH/3T3 cells. IP: HeLa, NIH/3T3 cells.
General notes
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.

Form

Liquid
Storage instructions

Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
Storage buffer

pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59.94% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR24997-22
Isotype

IgG
Research areas

Alternative Versions
- Anti-Calcineurin A antibody [EPR24997-22] - BSA and Azide free (ab282152)
Compatible Secondaries
Dyes/Markers
- Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (ab195889)
Isotype control
- Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
Related Buffer
- Antigen Retrieval Buffer (100X Tris-EDTA Buffer, pH 9.0) (ab93684)
Related Products
- Anti-GAPDH antibody [6C5] - Loading Control (ab8245)

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab282104 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
IP		1/30.
ICC/IF		1/50.
IHC-P		1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Flow Cyt (Intra)		1/500.
WB		1/1000. Predicted molecular weight: 59 kDa.

Notes
IP 1/30.
ICC/IF 1/50.
IHC-P 1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Flow Cyt (Intra) 1/500.
WB 1/1000. Predicted molecular weight: 59 kDa.

Function

Calcium-dependent, calmodulin-stimulated protein phosphatase. This subunit may have a role in the calmodulin activation of calcineurin. Dephosphorylates DNM1L, HSPB1 and SSH1.
Sequence similarities

Belongs to the PPP phosphatase family. PP-2B subfamily.
Cellular localization

Nucleus. Colocalizes with ACTN1 and MYOZ2 at the Z line in heart and skeletal muscle.
Target information above from: UniProt accession Q08209 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 5530 Human
- Entrez Gene: 19055 Mouse
- Entrez Gene: 24674 Rat
- Omim: 114105 Human
- SwissProt: Q08209 Human
- SwissProt: P63328 Mouse
- SwissProt: P63329 Rat
- Unigene: 435512 Human
- Unigene: 331389 Mouse
- Unigene: 6866 Rat
see all
Alternative names
- Alpha isoform formerly PPP2B antibody
- Calcineurin A alpha antibody
- Calcineurin A1 antibody
- CalcineurinA antibody
- Calmodulin dependent calcineurin A subunit alpha isoform antibody
- Calmodulin-dependent calcineurin A subunit alpha isoform antibody
- CALN antibody
- CALNA 1 antibody
- CALNA antibody
- CALNA1 antibody
- CAM PRP catalytic subunit antibody
- CAM-PRP catalytic subunit antibody
- CCN 1 antibody
- CCN1 antibody
- CNA 1 antibody
- CNA alpha antibody
- CNA antibody
- CNA1 antibody
- PP2BA_HUMAN antibody
- PPP2B antibody
- Ppp3ca antibody
- Protein phosphatase 2B catalytic subunit antibody
- Protein phosphatase 3 (formerly 2B) catalytic subunit alpha isoform antibody
- Protein phosphatase 3 catalytic subunit alpha isoform PPP3CA antibody
- Protein phosphatase 3 catalytic subunit alpha isozyme antibody
- Serine/threonine protein phosphatase 2B catalytic subunit alpha isoform antibody
- Serine/threonine-protein phosphatase 2B catalytic subunit alpha isoform antibody
see all

Western blot - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

All lanes : Anti-Calcineurin A antibody [EPR24997-22] (ab282104) at 1/1000 dilution

Lane 1 : Wild-type HAP1 (human chronic myelogenous leukemia near-haploid cell) whole cell lysate
Lane 2 : PPP3CA knockout HAP1 (human chronic myelogenous leukemia near-haploid cell) whole cell lysate
Lane 3 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
Lane 4 : SH-SY5Y (human neuroblastoma epithelial cell) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (IRDye® 800CW) (ab216773) and Goat Anti-Mouse IgG H&L (IRDye® 680RD) (ab216776) at 1/10000 dilution

Predicted band size: 59 kDa
Observed band size: 32,59 kDa why is the actual band size different from the predicted?

Blocking and diluting buffer and concentration: Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS

Lanes 1-4: Merged signal (red and green). Green - ab282104 observed at 59/32 kDa. Red - loading control ab8245 observed at 36 kDa.

ab282104 Anti-PPP3CA antibody [EPR24997-22 HT] was shown to specifically react with PPP3CA in wild-type HAP1 cells. Loss of signal was observed when the knockout cell line was used. Wild-type and PPP3CA knockout samples were subjected to SDS-PAGE. ab282104 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at 4? overnight at 1/1000 dilution and 1/20000 dilution respectively. 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 1/10000 dilution for 1 hour at room temperature before imaging.

The expression pattern & observed MW are consistent with what has been described in the literature (PMID:17785681).
Western blot - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

All lanes : Anti-Calcineurin A antibody [EPR24997-22] (ab282104) at 1/1000 dilution

Lane 1 : Mouse brain tissue lysate
Lane 2 : Rat brain tissue lysate
Lane 3 : Mouse lung tissue lysate
Lane 4 : Rat lung tissue lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)

Predicted band size: 59 kDa
Observed band size: 32,59 kDa why is the actual band size different from the predicted?

Blocking and diluting buffer and concentration: 5% NFDM/TBST

The expression pattern & observed MW are consistent with what has been described in the literature (PMID: 17785681).

Exposure time: Lane 1-2: 3.25 seconds Lane 3: 26 seconds Lane 4: 59 seconds
Western blot - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

All lanes : Anti-Calcineurin A antibody [EPR24997-22] (ab282104) at 1/1000 dilution

Lane 1 : Human PPP3CA recombinant protein
Lane 2 : Human PPP3CB recombinant protein
Lane 3 : Human PPP3CC recombinant protein

Lysates/proteins at 0.01 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 59 kDa
Observed band size: 32,59 kDa why is the actual band size different from the predicted?

Blocking and diluting buffer and concentration: 5% NFDM/TBST

This antibody does not recognize human PPP3CB and PPP3CC. These rec proteins are made in-house and expressed from E.coli expression systems.

Exposure time: 15 seconds
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Immunohistochemical analysis of paraffin-embedded Human cerebrum tissue labelling Calcineurin A with ab282104 at 1/2000 dilution (0.267 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Cytoplasmic staining on human cerebrum (PMID: 28524178). The section was incubated with ab282104 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Immunohistochemical analysis of paraffin-embedded Human astrocytoma tissue labelling Calcineurin A with ab282104 at 1/2000 dilution (0.267 ug/ml) followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer). Cytoplamic staining on human astrocytoma. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer).

Heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Immunohistochemical analysis of paraffin-embedded Human colon carcinoma tissue labelling Calcineurin A with ab282104 at 1/2000 dilution (0.267 ug/ml) followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer). Cytoplamic staining on human colon carcinoma. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer).

Heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labelling Calcineurin A with ab282104 at 1/2000 dilution (0.267 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Cytoplasmic staining on mouse cerebrum. The section was incubated with ab282104 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Immunohistochemical analysis of paraffin-embedded Rat cerebrum tissue labelling Calcineurin A with ab282104 at 1/2000 dilution (0.267 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Cytoplasmic staining on rat cerebrum. The section was incubated with ab282104 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Immunohistochemical analysis of paraffin-embedded Parental HAP1 cell pellet (A) and PPP3CA KO HAP1 cell pellet (B) labelling Calcineurin A with ab282104 at 1/100 dilution (5.33 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Cytoplasmic staining on parental HAP1 cell pellet (A) and no staining on PPP3CA KO HAP1 cell pellet (B). The section was incubated with ab282104 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins
Immunocytochemistry/ Immunofluorescence - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized HeLa (Human cervix adenocarcinoma) cells labelling Calcineurin A with ab282104 at 1/50 dilution (10.66 ug/ml), followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in HeLa cell line is observed.

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution.
Immunocytochemistry/ Immunofluorescence - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized NIH/3T3 (Mouse embryonic fibroblast) cells labelling Calcineurin A with ab282104 at 1/50 dilution (10.66 ug/ml), followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in NIH/3T3 cell line is observed.

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution.
Flow Cytometry (Intracellular) - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa (Human cervix adenocarcinoma) cells labelling Calcineurin A with ab282104 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
Flow Cytometry (Intracellular) - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized NIH/3T3 (Mouse embryonic fibroblast) cells labelling Calcineurin A with ab282104 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
Immunoprecipitation - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Calcineurin A was immunoprecipitated from 0.35 mg HeLa (Human cervix adenocarcinoma) whole cell lysate with ab282104 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab282104 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1: HeLa (Human cervix adenocarcinoma) whole cell lysate 10 ug

Lane 2: ab282104 IP in HeLa whole cell lysate

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab282104 in HeLa whole cell lysate

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 3 minutes
Immunoprecipitation - Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

Calcineurin A was immunoprecipitated from 0.35 mg NIH/3T3 (mouse embryonic fibroblast) whole cell lysate with ab282104 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab282104 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1: NIH/3T3 (mouse embryonic fibroblast) whole cell lysate10 ug

Lane 2: ab282104 IP in NIH/3T3 whole cell lysate

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab282104 in NIH/3T3 whole cell lysate

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 59 seconds
Anti-Calcineurin A antibody [EPR24997-22] (ab282104)

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

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Key features and details

Related conjugates and formulations

Overview

Product name

Description

Host species

Tested applications

Species reactivity

Immunogen

Positive control

General notes

Properties

Form

Storage instructions

Storage buffer

Purity

Clonality

Clone number

Isotype

Research areas

Associated products

Alternative Versions

Compatible Secondaries

Dyes/Markers

Isotype control

Related Buffer

Related Products

Applications

The Abpromise guarantee

Target

Function

Sequence similarities

Cellular localization

Database links

Alternative names

Images

Protocols

Datasheets and documents

SDS download

Datasheet download

Certificate of Compliance

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