Anti-CrkL antibody [Y244]

7 product images

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  Flow Cytometry - Anti-CrkL antibody [Y244] (ab32018)

  Flow Cytometry analysis of K-562 (Human chronic myelogenous leukemia lymphoblast) cells labelling CrkL with purified ab32018 at 1/20 dilution (10 µg/mL) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabelled control - Cell without incubation with primary antibody and secondary antibody (Blue).

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  Western blot - Anti-CrkL antibody [Y244] (ab32018)

  All lanes: Western blot - Anti-CrkL antibody [Y244] (ab32018) at 1/1000 dilution

  Lane 1: K-562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysate

  Lane 2: Mouse thymus lysate

  Lane 3: Rat thymus lysate

  Secondary

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

  Predicted band size: 34 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CrkL antibody [Y244] (ab32018)

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human colon tissue sections labeling CrkL with purified ab32018 at 1/100 dilution (2.24 μg/mL). Heat mediated antigen retrieval was performed using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Tissue was counterstained with Hematoxylin. Rabbit specific IHC polymer detection kit HRP/DAB (Rabbit specific IHC polymer detection kit HRP/DAB ab209101) secondary antibody was used at 1/0 dilution. PBS instead of the primary antibody was used as the negative control.
  

  The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

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  Immunoprecipitation - Anti-CrkL antibody [Y244] (ab32018)

  Purified ab32018 at 1/50 dilution (2μg) immunoprecipitating CrkL in K-562 whole cell lysate.
  Lane 1 (input): K-562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysate 10μg
  Lane 2 (+): ab32018 + K-562 whole cell lysate.
  Lane 3 (-): Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab32018 in K-562 whole cell lysate.
  VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) (1/1000 dilution) was used for Western blotting.
  Blocking Buffer and concentration: 5% NFDM/TBST.
  Diluting buffer and concentration: 5% NFDM/TBST.
  Observed band size: 37 kDa

  All lanes: Immunoprecipitation - Anti-CrkL antibody [Y244] (ab32018)

  Predicted band size: 34 kDa

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  Western blot - Anti-CrkL antibody [Y244] (ab32018)

  Lanes 1-3: Merged signal (red and green). Green - ab32018 observed at 37 kDa. Red - loading control Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291 observed at 50 kDa.
  

  ab32018 Anti-CrkL antibody [Y244] was shown to specifically react with CrkL in wild-type HeLa cells. Loss of signal was observed when knockout cell line Human CRKL knockout HeLa cell line ab265993 (knockout cell lysate Human CRKL knockout HeLa cell lysate ab257397) was used. Wild-type and CrkL knockout samples were subjected to SDS-PAGE. ab32018 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (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.

  All lanes: Western blot - Anti-CrkL antibody [Y244] (ab32018) at 1/1000 dilution

  Lane 1: Wild-type HeLa cell lysate at 20 µg

  Lane 2: CRKL knockout HeLa cell lysate at 20 µg

  Lane 2: Western blot - Human CRKL knockout HeLa cell line (Human CRKL knockout HeLa cell line ab265993)

  Lane 3: K-562 cell lysate at 20 µg

  Secondary

  All lanes: Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) at 1/10000 dilution

  Predicted band size: 34 kDa

  Observed band size: 37 kDa

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  Immunocytochemistry/ Immunofluorescence - Anti-CrkL antibody [Y244] (ab32018)

  Immunocytochemistry analysis of K-562 (Human chronic myelogenous leukemia lymphoblast) cells labeling CrkL with purified ab32018 at 1/50 dilution (4.5 μg/mL). Cells were fixed in 100% Methanol and permeabilized with 0.1% tritonX-100. Cells were counterstained with Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1/200 (2.5 μg/mL). Goat anti rabbit IgG (Alexa Fluor^®488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) was used as the secondary antibody at 1/1000 (2 μg/mL) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

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  Image collected and cropped by CiteAb under a CC-BY license from the publication

  Western blot - Anti-CrkL antibody [Y244] (ab32018)

  CrkL western blot using anti-CrkL antibody [Y244] ab32018. Publication image and figure legend from Kostrzewska-Poczekaj, M., Bednarek, K., et al., 2020, Sci Rep, PubMed 31913340.

  
  

  ab32018 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab32018 please see the product overview.

  SiRNA mediated CRKL knockdown in UT-SCC-6A, UT-SCC-11 and UT-SCC-29 cell lines (RT-qPCR and Western Blot).