Recombinant Anti-PAK1 antibody [EPR20048] (ab223849)
- Datasheet
- References
- Protocols
Overview
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Product nameAnti-PAK1 antibody [EPR20048]
See all PAK1 primary antibodies -
DescriptionRabbit monoclonal [EPR20048] to PAK1
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Host speciesRabbit
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Tested applicationsSuitable for: WB, ICC/IF, Flow Cyt, IPmore details
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Species reactivityReacts with: Mouse, Rat, Human
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Immunogen
Synthetic peptide within Human PAK1 aa 200-300. The exact sequence is proprietary.
Database link: Q13153 -
Positive control
- WB: His-tagged human PAK1 (aa1-250) recombinant protein; SK-OV-3, SH-SY5Y, HEK-293T, NIH/3T3 and PC-12 whole cell lysates; Human fetal brain lysate; Mouse and rat brain lysates. ICC/IF: SH-SY5Y and HeLa cells. Flow Cyt: SH-SY5Y and HeLa cells. IP: SH-SY5Y whole cell lysate.
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General notes
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.
This product is a recombinant rabbit monoclonal antibody.
Properties
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FormLiquid
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Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
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Storage bufferPreservative: 0.1% Sodium azide
Constituents: 0.05% BSA, 40% Glycerol, PBS -
Concentration information loading... -
PurityProtein A purified
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ClonalityMonoclonal
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Clone numberEPR20048
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IsotypeIgG
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Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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Recombinant Protein
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Related Products
Applications
Our Abpromise guarantee covers the use of ab223849 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 |
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| WB | 1/1000. Detects a band of approximately 60 kDa (predicted molecular weight: 61 kDa). In WB this antibody showed weak staining of PAK1 on HeLa cell lysate |
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| ICC/IF | 1/100. | |
| Flow Cyt | 1/50. | |
| IP | 1/30. |
Target
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FunctionThe activated kinase acts on a variety of targets. Likely to be the GTPase effector that links the Rho-related GTPases to the JNK MAP kinase pathway. Activated by CDC42 and RAC1. Involved in dissolution of stress fibers and reorganization of focal complexes. Involved in regulation of microtubule biogenesis through phosphorylation of TBCB. Activity is inhibited in cells undergoing apoptosis, potentially due to binding of CDC2L1 and CDC2L2.
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Sequence similaritiesBelongs to the protein kinase superfamily. STE Ser/Thr protein kinase family. STE20 subfamily.
Contains 1 CRIB domain.
Contains 1 protein kinase domain. -
Post-translational
modificationsAutophosphorylated when activated by CDC42/p21 and RAC1. -
Cellular localizationCytoplasm. Cell junction > focal adhesion. Recruited to focal adhesions upon activation.
- Information by UniProt
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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
see all -
Alternative names
- ADRB2 antibody
- Alpha PAK antibody
- Alpha-PAK antibody
see all
Images
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![Western blot - Anti-PAK1 antibody [EPR20048] (ab223849)](https://www.abcam.com/ps/products/223/ab223849/Images/ab223849-309606-ab223849KOWBMPES.jpg)
Western blot - Anti-PAK1 antibody [EPR20048] (ab223849)All lanes : Anti-PAK1 antibody [EPR20048] (ab223849) at 1/1000 dilution
Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : PAK1 knockout HAP1 whole cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 61 kDaLanes 1 - 2: Merged signal (red and green). Green - ab223849 observed at 61 kDa. Red - loading control, ab9484, observed at 37 kDa.
ab223849 was shown to specifically react with PAK1 in wild-type HAP1 cells as signal was lost in PAK1 knockout cells. Wild-type and PAK1 knockout samples were subjected to SDS-PAGE. Ab223849 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
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![Western blot - Anti-PAK1 antibody [EPR20048] (ab223849)](https://www.abcam.com/ps/products/223/ab223849/Images/ab223849-286960-2238497.jpg)
Western blot - Anti-PAK1 antibody [EPR20048] (ab223849)All lanes : Anti-PAK1 antibody [EPR20048] (ab223849) at 1/1000 dilution
Lane 1 : SK-OV-3 (human ovarian cancer epithelial cell line) whole cell lysate
Lane 2 : NIH/3T3 (mouse embryo fibroblast cell line) whole cell lysate
Lane 3 : PC-12 (rat adrenal gland pheochromocytoma cell line) whole cell lysate
Lane 4 : Human fetal brain lysate
Lysates/proteins at 20 µg per lane.
Secondary
Lanes 1-3 : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Lane 4 : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/4000 dilution
Developed using the ECL technique.
Predicted band size: 61 kDa
Observed band size: 60 kDa why is the actual band size different from the predicted?Exposure time : Lane 1: 3 minutes; Lane 2: 30 seconds; Lane 3: 3 seconds; Lane 4: 15 seconds.
Blocking/Dilution buffer: 5% NFDM/TBST.
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![Immunocytochemistry/ Immunofluorescence - Anti-PAK1 antibody [EPR20048] (ab223849)](https://www.abcam.com/ps/products/223/ab223849/Images/ab223849-286958-2238495.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-PAK1 antibody [EPR20048] (ab223849)Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized SH-SY5Y (human neuroblastoma cell line from bone marrow) cells labeling PAK1 with ab223849 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on SH-SY5Y cell line.
The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.
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![Flow Cytometry - Anti-PAK1 antibody [EPR20048] (ab223849)](https://www.abcam.com/ps/products/223/ab223849/Images/ab223849-286956-2238493.jpg)
Flow Cytometry - Anti-PAK1 antibody [EPR20048] (ab223849)Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized SH-SY5Y (human neuroblastoma cell line from bone marrow) cell line labeling PAK1 with ab223849 at 1/500 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) at 1/2000 dilution was used as the secondary
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![Immunoprecipitation - Anti-PAK1 antibody [EPR20048] (ab223849)](https://www.abcam.com/ps/products/223/ab223849/Images/ab223849-286954-2238491.jpg)
Immunoprecipitation - Anti-PAK1 antibody [EPR20048] (ab223849)PAK1 was immunoprecipitated from 0.35 mg SH-SY5Y (human neuroblastoma cell line from bone marrow) whole cell lysate with ab223849 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab223849 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.
Lane 1: SH-SY5Y whole cell lysate 10 μg (Input).
Lane 2: ab223849 IP in SH-SY5Y whole cell lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab223849 in SH-SY5Y whole cell lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 1 second.
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![Western blot - Anti-PAK1 antibody [EPR20048] (ab223849)](https://www.abcam.com/ps/products/223/ab223849/Images/ab223849-286959-2238496.jpg)
Western blot - Anti-PAK1 antibody [EPR20048] (ab223849)All lanes : Anti-PAK1 antibody [EPR20048] (ab223849) at 1/1000 dilution
Lane 1 : Mouse brain lysate
Lane 2 : Rat brain lysate
Lane 3 : SH-SY5Y (human euroblastoma cell line from bone marrow) whole cell lysate
Lane 4 : HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Developed using the ECL technique.
Predicted band size: 61 kDa
Observed band size: 60 kDa why is the actual band size different from the predicted?Exposure time : Lane 1: 5 seconds; Lane 2: 3 seconds; Lane 3: 3 minutes; Lane 4: 1 minutes.
Blocking/Dilution buffer: 5% NFDM/TBST.
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![Immunocytochemistry/ Immunofluorescence - Anti-PAK1 antibody [EPR20048] (ab223849)](https://www.abcam.com/ps/products/223/ab223849/Images/ab223849-286957-2238494.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-PAK1 antibody [EPR20048] (ab223849)Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling PAK1 with ab223849 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on HeLa cell line.
The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.
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![Flow Cytometry - Anti-PAK1 antibody [EPR20048] (ab223849)](https://www.abcam.com/ps/products/223/ab223849/Images/ab223849-286955-2238492.jpg)
Flow Cytometry - Anti-PAK1 antibody [EPR20048] (ab223849)Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cell line labeling PAK1 with ab223849 at 1/50 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody.
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![Western blot - Anti-PAK1 antibody [EPR20048] (ab223849)](https://www.abcam.com/ps/products/223/ab223849/Images/ab223849-286961-2238498.jpg)
Western blot - Anti-PAK1 antibody [EPR20048] (ab223849)All lanes : Anti-PAK1 antibody [EPR20048] (ab223849) at 1/1000 dilution
Lane 1 : His-tagged human PAK1 (aa1-250) recombinant protein
Lane 2 : His-tagged human PAK2 (aa1-250) 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
Developed using the ECL technique.
Predicted band size: 61 kDa
Observed band size: 37 kDa why is the actual band size different from the predicted?Exposure time : Lane 1: 1 minute; Lane 2: 3 minutes.
Blocking/Dilution buffer: 5% NFDM/TBST.
References
ab223849 has not yet been referenced specifically in any publications.
