Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)

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Overview

  • Product name

    Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free
    See all Cdk4 primary antibodies

  • Description

    Rabbit monoclonal [EPR2513Y] to Cdk4 - BSA and Azide free

  • Host species

    Rabbit

  • Tested applications

    Suitable for: IP, ICC/IF, Flow Cyt, WBmore details

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Synthetic peptide corresponding to Human Cdk4 aa 1-100 (N terminal).

  • Positive control

    • WB: WT HAP1 whole cell lysate.

  • General notes

    Ab236042 is the carrier-free version of ab68266. 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 <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    ab236042 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

    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

Applications

Our Abpromise guarantee covers the use of ab236042 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
IP Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
WB Use at an assay dependent concentration. Detects a band of approximately 34 kDa (predicted molecular weight: 34 kDa).

Target

  • Function

    Ser/Thr-kinase component of cyclin D-CDK4 (DC) complexes that phosphorylate and inhibit members of the retinoblastoma (RB) protein family including RB1 and regulate the cell-cycle during G(1)/S transition. Phosphorylation of RB1 allows dissociation of the transcription factor E2F from the RB/E2F complexes and the subsequent transcription of E2F target genes which are responsible for the progression through the G(1) phase. Hypophosphorylates RB1 in early G(1) phase. Cyclin D-CDK4 complexes are major integrators of various mitogenenic and antimitogenic signals. Also phosphorylates SMAD3 in a cell-cycle-dependent manner and represses its transcriptional activity. Component of the ternary complex, cyclin D/CDK4/CDKN1B, required for nuclear translocation and activity of the cyclin D-CDK4 complex.

  • Involvement in disease

    Defects in CDK4 are a cause of susceptibility to cutaneous malignant melanoma type 3 (CMM3) [MIM:609048]. Malignant melanoma is a malignant neoplasm of melanocytes, arising de novo or from a pre-existing benign nevus, which occurs most often in the skin but also may involve other sites.

  • Sequence similarities

    Belongs to the protein kinase superfamily. CMGC Ser/Thr protein kinase family. CDC2/CDKX subfamily.
    Contains 1 protein kinase domain.

  • Post-translational
    modifications

    Phosphorylation at Thr-172 is required for enzymatic activity. Phosphorylated, in vitro, at this site by CCNH-CDK7, but, in vivo, appears to be phosphorylated by a proline-directed kinase. In the cyclin D-CDK4-CDKN1B complex, this phosphorylation and consequent CDK4 enzyme activity, is dependent on the tyrosine phosphorylation state of CDKN1B. Thus, in proliferating cells, CDK4 within the complex is phosphorylated on Thr-172 in the T-loop. In resting cells, phosphorylation on Thr-172 is prevented by the non-tyrosine-phosphorylated form of CDKN1B.

  • Cellular localization

    Cytoplasm. Nucleus. Membrane. Cytoplasmic when non-complexed. Forms a cyclin D-CDK4 complex in the cytoplasm as cells progress through G(1) phase. The complex accumulates on the nuclear membrane and enters the nucleus on transition from G(1) to S phase. Also present in nucleoli and heterochromatin lumps. Colocalizes with RB1 after release into the nucleus.
  • Information by UniProt

  • Database links

    see all

  • Alternative names

    • Cdk 4 antibody
    • cdk4 antibody
    • CDK4 protein antibody
    • CDK4_HUMAN antibody
    • Cell division kinase 4 antibody
    • Cell division protein kinase 4 antibody
    • CMM 3 antibody
    • CMM3 antibody
    • Crk3 antibody
    • Cyclin dependent kinase 4 antibody
    • Cyclin-dependent kinase 4 antibody
    • Melanoma cutaneous malignant 3 antibody
    • MGC14458 antibody
    • p34 cdk4 antibody
    • PSK J3 antibody
    • PSK-J3 antibody
    see all

Images

  • Western blot - Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)
    Western blot - Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)

    Lanes 1, 3 and 5: Wild-type HAP1 cell lysate (20 µg)
    Lanes 2, 4 and 6: CDK4 knockout HAP1 cell lysate (20 µg)
    Lanes 1 and 2: Green signal from target - ab68266 observed at 34 kDa 
    Lanes 3 and 4: Red signal from loading control - ab8226 observed at 42 kDa
    Lanes 5 and 6: Merged (red and green) signal 
    Unpurified ab68266 was shown to specifically react with CDK4 when CDK4 knockout samples were used. Wild-type and CDK4 knockout samples were subjected to SDS-PAGE. ab68266 and ab8226 (loading control to beta actin) were both diluted at 1/1000 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 1/10000 dilution for 1 h at room temperature before imaging.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab68266).

  • Immunoprecipitation - Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)
    Immunoprecipitation - Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)

    ab68266 (purified) at 1:30 dilution (2µg) immunoprecipitating Cdk4 in K562 whole cell lysate.
    Lane 1 (input): K562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysate 10µg
    Lane 2 (+): ab68266 & K562 whole cell lysate
    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab68266 in K562 whole cell lysate
    For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1:1000 dilution.
    Blocking and diluting buffer: 5% NFDM/TBST.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab68266).

  • Flow Cytometry - Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)
    Flow Cytometry - Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)
    Flow Cytometry analysis of A549 (Human lung carcinoma epithelial cell) cells labeling Cdk4 with purified ab68266 at 1:50 dilution (1 µg/ml) (red). Cells were fixed with 4% Paraformaldehyde. A Goat anti rabbit IgG (Alexa Fluor® 488) secondary antibody was used at 1:2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab68266).

  • Immunocytochemistry/ Immunofluorescence - Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)
    Immunocytochemistry/ Immunofluorescence - Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)
    Immunocytochemistry/ Immunofluorescence analysis of MCF7 (Human breast adenocarcinoma epithelial cell) cells labeling Cdk4 with Purified ab68266 at 1:100 (5.4 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with None. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1:1000 (2 ?g/ml) dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab68266).

  • Immunocytochemistry/ Immunofluorescence - Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)
    Immunocytochemistry/ Immunofluorescence - Anti-Cdk4 antibody [EPR2513Y] - BSA and Azide free (ab236042)

    Unpurified ab68266 staining Cdk4 in wild-type HAP1 cells (top panel) and Cdk4 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 ab68266 at 1/250 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 (ab68266).

References

ab236042 has not yet been referenced specifically in any publications.

Publishing research using ab236042? Please let us know so that we can cite the reference in this datasheet.

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