Astana Biomed Group, an authorized Abcam distributor in Central Asia

Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR4515] to Cdk6 - BSA and Azide free
  • Suitable for: ICC, Flow Cyt, WB, IHC-P
  • Knockout validated
  • Reacts with: Human

Overview

  • Product name

    Anti-Cdk6 antibody [EPR4515] - BSA and Azide free
    See all Cdk6 primary antibodies

  • Description

    Rabbit monoclonal [EPR4515] to Cdk6 - BSA and Azide free

  • Host species

    Rabbit

  • Tested applications

    Suitable for: ICC, Flow Cyt, WB, IHC-Pmore details
    Unsuitable for: IP

  • Species reactivity

    Reacts with: Human

  • Immunogen

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

  • Positive control

    • WB: Jurkat, K562, HeLa, HAP1, and 293T cell lysates. IHC-P:Human tonsil tissue. ICC/IF: HeLa and HAP1 cells.

  • General notes

    ab222395 is the carrier-free version of ab124821. 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

    Ab222395 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

    Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

    Reproducibility is key to advancing scientific discovery and accelerating scientists’ next breakthrough.

    Abcam is leading the way with our range of recombinant antibodies, knockout-validated antibodies and knockout cell lines, all of which support improved reproducibility.

    We are also planning to innovate the way in which we present recommended applications and species on our product datasheets, so that only applications & species that have been tested in our own labs, our suppliers or by selected trusted collaborators are covered by our Abpromise™ guarantee.

    In preparation for this, we have started to update the applications & species that this product is Abpromise guaranteed for.

    We are also updating the applications & species that this product has been “predicted to work with,” however this information is not covered by our Abpromise guarantee.

    Applications & species from publications and Abreviews that have not been tested in our own labs or in those of our suppliers are not covered by the Abpromise guarantee.

    Please check that this product meets your needs before purchasing. If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, as well as customer reviews and Q&As.

Properties

Images

  • Western blot - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Western blot - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    All lanes : Anti-Cdk6 antibody [EPR4515] (ab124821) at 1/1000 dilution

    Lane 1 : Wild-type HeLa cell lysate
    Lane 2 : CDK6 knockout HeLa cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 37 kDa
    Observed band size: 37 kDa



    This data was developed using the same antibody clone in a different buffer formulation (ab124821).

      Lanes 1- 2: Merged signal (red and green). Green - ab124821 observed at 37 kDa. Red - Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) observed at 50 kDa.

     ab124821 was shown to react with Cdk6 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab266059 (knockout cell lysate ab257088) was used. Wild-type HeLa and CDK6 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. ab124821 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) 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 - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Immunocytochemistry - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)

    ab124821 staining Cdk6 in wild-type HAP1 cells (top panel) and Cdk6 knockout HAP1 cells (bottom panel). The cells were fixed with 4% formaldehyde (10min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab124821 at 1/500 dilution and ab195889 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab124821).

  • Flow Cytometry - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Flow Cytometry - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Overlay histogram showing HeLa cells stained with ab124821 (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 (ab124821, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1µg/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.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab124821).

  • Western blot - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Western blot - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Lanes 1-2 : Anti-Cdk6 antibody [EPR4515] (ab124821) at 1/10000 dilution
    Lanes 3-4 : Anti-beta Actin antibody [mAbcam 8226] - Loading Control (ab8226) at 1/1000 dilution


    Lanes 1 & 3 & 5 : Wild-type HAP1 cell lysate
    Lanes 2 & 4 & 6 : CDK6 knockout HAP1 cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 37 kDa



    Lanes 1 and 2: Green signal from target - ab124821 observed at 37 kDa 
    Lanes 3 and 4: Red signal from loading control - ab8226 observed at 42 kDa
    Lanes 5 and 6: Merged (red and green) signal 
    ab124821 was shown to specifically react with CDK6 when CDK6 knockout samples were used. Wild-type and CDK6 knockout samples were subjected to SDS-PAGE. ab124821 and ab8226 (loading control to beta actin) were diluted at 1/10 000 and 1/1000 respectively and incubated overnight at 4°C. 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 1/10,000 dilution for 1 h at room temperature before imaging.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)

    ab124821 at 1/100 dilution, staining Cdk6 in formalin-fixed paraffin-embedded human tonsil tissue by immunohistochemistry.

    Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab124821).

  • Immunocytochemistry - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Immunocytochemistry - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)

    Clone EPR4515 (ab222395) has been successfully conjugated by Abcam. This image was generated using Anti-Cdk6 antibody [EPR4515] (Alexa Fluor® 647). Please refer to ab198946 for protocol details.

    ab198946 staining Cdk6 in HeLa cells. The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab198946 at 1/100 dilution (shown in red) and ab195887, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 488), at 1/250 dilution (shown in green). Nuclear DNA was labelled with DAPI (shown in blue).

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

    This product also gave a positive signal under the same testing conditions in HeLa cells fixed with 100% methanol (5min).

  • Immunocytochemistry - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Immunocytochemistry - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)

    Clone EPR4515 (ab222395) has been successfully conjugated by Abcam. This image was generated using Anti-Cdk6 antibody [EPR4515] (Alexa Fluor® 488). Please refer to ab198944 for protocol details.

    ab198944 staining Cdk6 in HeLa cells. The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab198944 at 1/100 dilution (shown in green) and ab7291 (Mouse monoclonal [DM1A] to alpha Tubulin) at 1µg/ml. This was followed by an incubation at room temperature for 1h with ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor® 594), pre-adsorbed, at 1µg/ml (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • Immunocytochemistry - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Immunocytochemistry - Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)

    ICC/IF image of ab124821 at 1/100 dilution, staining Cdk6 in HeLa cells.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab124821).

  • Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)
    Anti-Cdk6 antibody [EPR4515] - BSA and Azide free (ab222395)

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