Name: Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y]
Brand: RabMAb
SKU: ab40795
Rating: 4 (4 reviews)

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Western blot - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EP656Y] to PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154)
Suitable for: Flow Cyt (Intra), WB, IHC-P, IP, ICC/IF
Reacts with: Mouse, Rat, Human

Product name

Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y]
See all PAK1+PAK2+PAK3 primary antibodies
Description

Rabbit monoclonal [EP656Y] to PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154)
Host species

Rabbit
Specificity

ab203958 recognises p21-activated kinase 1 (PAK1).
Tested applications

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

Reacts with: Mouse, Rat, Human
Immunogen

Synthetic peptide within Human PAK1 (phospho S144). The exact sequence is proprietary.
Database link: Q13153
Positive control
- WB: MCF7, HeLa, RAW 264.7 and C6 cell lysates. IHC: Human liver carcinoma, mouse cerebral cortex, rat cerebral cortex. ICC/IF: HeLa cells. IP: HeLa cell lysate. Flow Cyt (intra): NIH/3T3 cell lysate.
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.
We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

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: 7.20
Preservative: 0.01% Sodium azide
Constituents: 40% Glycerol (glycerin, glycerine), 0.05% BSA, 59% PBS
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EP656Y
Isotype

IgG
Research areas

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab40795 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
Flow Cyt (Intra)		1/120. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
WB	(2)	1/10000 - 1/50000. Detects a band of approximately 66 kDa (predicted molecular weight: 65 kDa).
IHC-P	(1)	1/100 - 1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
IP		1/40.
ICC/IF	(1)	1/250 - 1/500.

Notes
Flow Cyt (Intra) 1/120. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
WB 1/10000 - 1/50000. Detects a band of approximately 66 kDa (predicted molecular weight: 65 kDa).
IHC-P 1/100 - 1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
IP 1/40.
ICC/IF 1/250 - 1/500.

Cellular localization

PAK1: Cytoplasm. Cell junction > focal adhesion. Recruited to focal adhesions upon activation. PAK1+PAK2+PAK3: PAK1: Cytoplasm. Cell junction > focal adhesion. Recruited to focal adhesions upon activation. PAK2: Cytoplasm and Nucleus. Cytoplasm > perinuclear region. Membrane. Interaction with ARHGAP10 probably changes PAK-2p34 location to cytoplasmic perinuclear region. Myristoylation changes PAK-2p34 location to the membrane. PAK3: Cytoplasmic
Database links
- Entrez Gene: 5058 Human
- Entrez Gene: 18479 Mouse
- Entrez Gene: 29431 Rat
- Omim: 602590 Human
- SwissProt: Q13153 Human
- SwissProt: O88643 Mouse
- SwissProt: P35465 Rat
- Unigene: 435714 Human
- Unigene: 260227 Mouse
- Unigene: 9149 Rat
see all

Western blot - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

All lanes : Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795) at 1/1000 dilution

Lane 1 : MCF7, grown in serum-free media overnight, whole cell lysate
Lane 2 : MCF7, grown in serum-free media overnight, then treated with EGF 1µg/ml for 10min, whole cell lysate
Lane 3 : MCF7, grown in serum-free media overnight, then treated with EGF 1µg/ml for 10min, whole cell lysate. The membrane was incubated with phosphatase.

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 65 kDa
Observed band size: 55 kDa why is the actual band size different from the predicted?

Exposure time: 1 minute

Blocking and dilution buffer: 5% NFDM/TBST.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

ab40795 staining PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) in rat cerebral cortex tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and antigen retrieval was by heat mediation in a EDTA buffer. Samples were incubated with primary antibody at a dilution of 1/100. A goat anti-rabbit IgG H&L (HRP) ab97051 was used as the secondary antibody at a dilution of 1/500.

Negative control 1: PBS in place of primary
Immunocytochemistry/ Immunofluorescence - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

ab40795 staining PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) in HeLa (human cervix adenocarcinoma) cells, treated and untreated with Lambda Protein Phosphtase 31℃ for 5h by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with 4% Paraformaldehyde and permeabilized with 0.1% Triton X-100. Samples were incubated with primary antibody at a dilution of 1/100. A goat anti rabbit IgG (Alexa Fluor® 488) (ab150077) was used as the secondary antibody. ab7291 and ab150120 were used as counterstains for primary antibody ab75748 and secondary antibody ab150077 respectively and DAPI was used as a nuclear counterstain.

Negative control 1: Rabbit primary antibody and anti-mouse secondary antibody (ab150120)
Negative control 2: Mouse primary antibody (ab7291) and anti-rabbit secondary antibody (ab150077)
Flow Cytometry (Intracellular) - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

ab40795 staining PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) in the human cell line NIH/3T3 (mouse embryo) by intracellular flow cytometry. Cells were fixed with 4% paraformaldehyde and the sample was incubated with the primary antibody at a dilution of 1/120. A goat anti rabbit IgG (Alexa Fluorr^® 488) at a dilution of 1/500 was used as the secondary antibody.

Isoytype control: Rabbit monoclonal IgG (Black)

Unlabelled control: Cell without incubation with primary antibody and secondary antibody (Blue)
Immunoprecipitation - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

ab40795 immunoprecipitating PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154). 10µg of HeLa (human cervix adenocarcinoma) whole cell lysate was incubated with primary antibody at a dilution of 1/40 and VeriBlot for IP Detection Reagent (HRP) (ab131366) at a dilution of 1/10000.

Lane 1: HeLa whole cell lysate (10ug)
Lane 2: ab40795 IP in HeLa whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab40795 in HeLa (human cervix adenocarcinoma) whole cell lysate
Western blot - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

All lanes : Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795) at 1/2000 dilution

Lane 1 : HeLa cell lysate with None
Lane 2 : HeLa cell lysate with PAK2 (pS141)
Lane 3 : HeLa cell lysate with PAK2 non-phospho
Lane 4 : HeLa cell lysate with PAK3 (pS154)
Lane 5 : HeLa cell lysate with PAK3 non-phospho

Predicted band size: 65 kDa
Western blot - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

All lanes : Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795) at 1/50000 dilution

Lane 1 : C6 (rat glioma) whole cell lysate - treated with phosphatase
Lane 2 : C6 (rat glioma) whole cell lysate - untreated

Lysates/proteins at 10 µg per lane.

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

Predicted band size: 65 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

ab40795 staining PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) in mouse cerebral cortex tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and antigen retrieval was by heat mediation in a EDTA buffer. Samples were incubated with primary antibody at a dilution of 1/100. A goat anti-rabbit IgG H&L (HRP) ab97051 was used as the secondary antibody at a dilution of 1/500.

Negative control 1: PBS in place of primary antibody.
Flow Cytometry (Intracellular) - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

Overlay histogram showing HeLa cells stained with unpurified ab40795 (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 (ab40795, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was DyLight^® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (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.
Western blot - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

All lanes : Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795) at 1/10000 dilution

Lane 1 : HeLa whole cell lysate - untreated
Lane 2 : HeLa whole cell lysate - treated with phosphatase

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 65 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

ab40795 staining PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) in human liver carcinoma tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and antigen retrieval was by heat mediation in a EDTA buffer. Samples were incubated with primary antibody at a dilution of 1/100. A goat anti-rabbit IgG H&L (HRP) ab97051 was used as the secondary antibody at a dilution of 1/500.

Negative control 1: PBS in place of primary antibody.
Western blot - Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

All lanes : Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795) at 1/10000 dilution

Lane 1 : RAW264.7 (mouse abelson murine leukemia virus-induced tumor) whole cell lysate - treated with phosphatase
Lane 2 : RAW264.7 (mouse abelson murine leukemia virus-induced tumor) whole cell lysate - untreated

Lysates/proteins at 10 µg per lane.

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

Predicted band size: 65 kDa
Anti-PAK1 (phospho S144) + PAK2 (phospho S141) + PAK3 (phospho S154) antibody [EP656Y] (ab40795)

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

SDS download

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Datasheet download

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Publishing research using ab40795? Please let us know so that we can cite the reference in this datasheet.

ab40795 has been referenced in 31 publications.

Kuželová K et al. Group I p21-activated kinases in leukemia cell adhesion to fibronectin. Cell Adh Migr 15:18-36 (2021). PubMed: 33464167
Tan S et al. RAC1 Involves in the Radioresistance by Mediating Epithelial-Mesenchymal Transition in Lung Cancer. Front Oncol 10:649 (2020). PubMed: 32411607
Schöttelndreier D et al. Low Density Lipoprotein Receptor-Related Protein-1 (LRP1) Is Involved in the Uptake of Clostridioides difficile Toxin A and Serves as an Internalizing Receptor. Front Cell Infect Microbiol 10:565465 (2020). PubMed: 33194803
Rodríguez-Fdez S et al. Vav2 catalysis-dependent pathways contribute to skeletal muscle growth and metabolic homeostasis. Nat Commun 11:5808 (2020). PubMed: 33199701
Zhang Y et al. PTPs inhibitors promote hematopoietic stem cell regeneration. Nat Commun 10:3667 (2019). PubMed: 31413255

View all Publications for this product

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1-4 of 4 Abreviews or Q&A

Application

Western blot

Sample

Human Cell lysate - whole cell (HCT116)

Loading amount

40 µg

Specification

HCT116

Gel Running Conditions

Non-reduced Denaturing (15%)

Blocking step

Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Nov 30 2012

Application

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)

Sample

Human Tissue sections (normal tissue array (including breast/colon/spleen)

Specification

normal tissue array (including breast/colon/spleen

Fixative

Formaldehyde

Antigen retrieval step

Heat mediated - Buffer/Enzyme Used: Citrate

Permeabilization

Blocking step

Serum as blocking agent for 10 minute(s) · Concentration: 10%

Abcam user community

Verified customer

Submitted May 19 2009

Application

Western blot

Sample

Human Cell lysate - whole cell (Breast)

Loading amount

20 µg

Specification

Breast

Gel Running Conditions

Non-reduced Non-Denaturing (Native) (4-12)

Blocking step

Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10%

Abcam user community

Verified customer

Submitted May 18 2009

Application

Immunocytochemistry/ Immunofluorescence

Sample

Mouse Cell (cultured hippocampal astrocytes)

Specification

cultured hippocampal astrocytes

Fixative

Paraformaldehyde

Blocking step

BSA as blocking agent for 20 minute(s) · Concentration: 1%

Abcam user community

Verified customer

Submitted Mar 08 2007

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Key features and details

Overview

Product name

Description

Host species

Specificity

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

Conjugation kits

Isotype control

Applications

The Abpromise guarantee

Target

Cellular localization

Database links

Images

Protocols

Datasheets and documents

SDS download

Datasheet download

References (31)

Customer reviews and Q&As

Western blot abreview for Anti-PAK1 (phospho S144) antibody [EP656Y]

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-PAK1 (phospho S144) antibody [EP656Y]

Western blot abreview for Anti-PAK1 (phospho S144) antibody [EP656Y]

Immunocytochemistry/ Immunofluorescence abreview for Anti-PAK1 (phospho S144) antibody [EP656Y]