Astana Biomed Group, an authorized Abcam distributor in Central Asia

Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR20048] to PAK1 - BSA and Azide free
  • Suitable for: Flow Cyt, ICC/IF, IP, WB
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Overview

  • Product name

    Anti-PAK1 antibody [EPR20048] - BSA and Azide free
    See all PAK1 primary antibodies

  • Description

    Rabbit monoclonal [EPR20048] to PAK1 - BSA and Azide free

  • Host species

    Rabbit

  • Tested Applications & Species

    Application Species
    Flow Cyt
    Human
    ICC/IF
    Human
    IP
    Human
    WB
    Human
    See all applications and species data

  • Immunogen

    Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • ICC/IF: SH-SY5Y and HeLa cells. Flow Cyt: SH-SY5Y and HeLa cells. IP: SH-SY5Y whole cell lysate. WB: HeLa and HAP1 cell lysates.

  • General notes

    Ab242421 is the carrier-free version of ab223849. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    Use our conjugation kits  for antibody conjugates that are ready-to-use in as little as 20 minutes with

    ab242421 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

    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.

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing the problem with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation.

    One factor contributing to the crisis is the use of antibodies that are not suitable. This can lead to misleading results and the use of incorrect data informing project assumptions and direction. To help address this challenge, we have introduced an application and species grid on our primary antibody datasheets to make it easy to simplify identification of the right antibody for your needs.

    Learn more here.

Properties

Images

  • Western blot - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)
    Western blot - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)
    All lanes : Anti-PAK1 antibody [EPR20048] (ab223849) at 1/1000 dilution

    Lane 1 : Wild-type HeLa cell lysate
    Lane 2 : PAK1 knockout HeLa cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 61 kDa
    Observed band size: 65 kDa
    why is the actual band size different from the predicted?



    This data was developed using the same antibody clone in a different buffer formulation (ab223849).

      Lanes 1- 2: Merged signal (red and green). Green - ab223849 observed at 65 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

     ab223849 was shown to react with PAK1 in wild-type HeLa cells in western blot. The band observed in knockout cell line ab264889 (knockout cell lysate ab257572) lane below 65kDa may represent truncated forms and cleaved fragments. This has not been investigated further. Wild-type HeLa and PAK1 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab223849 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at a 1 in 1000 dilution and a 1 in 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 in 20000 dilution for 1 hour at room temperature before imaging.

  • Immunocytochemistry/ Immunofluorescence - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)
    Immunocytochemistry/ Immunofluorescence - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)

    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.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab223849).

  • Flow Cytometry - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)
    Flow Cytometry - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)

    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 antibody.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab223849).

  • Immunoprecipitation - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)
    Immunoprecipitation - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)

    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.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab223849).

  • Western blot - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)
    Western blot - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)
    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 kDa



    Lanes 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.

    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.

  • Immunocytochemistry/ Immunofluorescence - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)
    Immunocytochemistry/ Immunofluorescence - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)

    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.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab223849).

  • Flow Cytometry - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)
    Flow Cytometry - Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)

    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.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab223849).

  • Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)
    Anti-PAK1 antibody [EPR20048] - BSA and Azide free (ab242421)

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