使用敲除细胞株进行验证RabMAb

Anti-Cdk2抗体[E304] (ab32147)

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  • 产品名称Anti-Cdk2抗体[E304]
    参阅全部 Cdk2 一抗
  • 描述
    兔单克隆抗体[E304] to Cdk2
  • 经测试应用适用于: ICC/IF, IP, WB, Flow Cyt, IHC-Pmore details
  • 种属反应性
    与反应: Mouse, Rat, Human
  • 免疫原

    A synthetic peptide corresponding to residues in C-terminus of human Cdk2

  • 表位The epitope is within the C-terminus of human Cdk2
  • 阳性对照
    • Hela cells Hela cell lysate.
  • 常规说明

    This product is a recombinant rabbit monoclonal antibody.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated ‘PUR’ on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    Produced using Abcam’s RabMAb® technology. RabMAb® technology is covered by the following U.S. Patents, No. 5,675,063 and/or 7,429,487.

    A trial size is available to purchase for this antibody.

     

    Alternative versions available:

    Anti-Cdk2 antibody (Alexa Fluor® 647) [E304] (ab206038)

    Anti-Cdk2 antibody (Alexa Fluor® 555) [E304] (ab208043)

    Anti-Cdk2 antibody (Alexa Fluor® 568) [E304] (ab208044)

性能

应用

Our Abpromise guarantee covers the use of ab32147 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

应用 Ab评论 说明
ICC/IF 1/200.

For unpurified use at 1/100.

IP 1/40.
WB 1/1000 - 1/10000. Detects a band of approximately 33 kDa (predicted molecular weight: 34 kDa).
Flow Cyt 1/80.

ab172730-Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

IHC-P 1/50. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

See protocols http://www.abcam.com/protocols/ihc-antigen-retrieval-protocol.

靶标

Anti-Cdk2 antibody [E304] 图像



  • Predicted band size : 34 kDa

    Lanes 1, 3 and 5: Wild-type HAP1 cell lysate (20 µg)
    Lanes 2, 4 and 6: CDK2 knockout HAP1 cell lysate (20 µg)
    Lanes 1 and 2: Green signal from target – ab32147 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

    ab32147 was shown to specifically react with CDK2 when CDK2 knockout samples were used. Wild-type and CDK2 knockout samples were subjected to SDS-PAGE. ab32147 and ab8226 (loading control to beta actin) were both diluted 1/1000 and incubated overnight at 4ºC. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) ab216776 secondary antibodies at 1/10 000 dilution for 1 h at room temperature before imaging.

  • ab32147 staining Cdk2 in Human tonsil tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed and paraffin-embedded, antigen retrieval was by heat mediation in Tris/EDTA buffer pH9. Samples were incubated with primary antibody (1/50). An undiluted HRP-conjugated mouse anti-rabbit IgG was used as the secondary antibody. Tissue counterstained with Hematoxylin. PBS was used in the negative control rather than the Primary antibody.

  • ab32147 staining Cdk2 in the HeLa cell line by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with 4% Paraformaldehyde permeabilized with 0.1% Triton X-100. Samples were incubated with primary antibody (1/200). ab150078(1/500) an Alexa Fluor®555-conjugated Goat anti-rabbit IgG was used as the secondary antibody. Nuclei were counterstained with DAPI.

  • Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labelling Cdk2 with purified ab32147 at 1/80 (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/2000) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

  • Immunofluorescent staining of HeLa cells using ab32147, unpurified.

  • ab32147 (purified) at 1/40 immunoprecipitating Cdk2 from HeLa cells(Lane 1). Lane 2 - PBS. For western blotting, a HRP-conjugated anti-rabbit IgG, specific to the non-reduced form of IgG was used as the secondary antibody (1/1000).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • ab32147 staining Cdk2 in Human squamouscell carcinoma of cervix tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed and paraffin-embedded, antigen retrieval was by heat mediation in Tris/EDTA buffer pH9. Samples were incubated with primary antibody (1/50). An undiluted HRP-conjugated mouse anti-rabbit IgG was used as the secondary antibody. Tissue counterstained with Hematoxylin. PBS was used in the negative control rather than the Primary antibody.

  • All lanes : Anti-Cdk2 antibody [E304] (ab32147) at 1/5000 dilution

    Lane 1 : C6 cell lysate
    Lane 2 : PC-12 cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG, (H+L), HRP-conjugated at 1/1000 dilution

    Predicted band size : 34 kDa
  • Anti-Cdk2 antibody [E304] (ab32147) at 1/1000 dilution + NIH/3T3 cell lysate at 20 µg

    Secondary
    Goat Anti-Rabbit IgG, (H+L), HRP-conjugated at 1/1000 dilution

    Predicted band size : 34 kDa
  • All lanes : Anti-Cdk2 antibody [E304] (ab32147) at 1/1000 dilution

    Lane 1 : Jurkat cell lysate
    Lane 2 : Hela cell lysate
    Lane 3 : K562 cell lysate
    Lane 4 : 293 cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG, (H+L), HRP-conjugated at 1/1000 dilution

    Predicted band size : 34 kDa
  • All lanes : Anti-Cdk2 antibody [E304] (ab32147) at 1/1000 dilution (unpurified)

    Lane 1 : Human osteosarcoma whole cell lysate - control, non-targeting siRNA
    Lane 2 : Human osteosarcoma whole cell lysate - siRNA for CDK2

    Lysates/proteins at 20 µg per lane.

    Secondary
    HRP-conjugated goat anti-rabbit IgG polyclonal at 1/2000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 34 kDa
    Observed band size : 34 kDa


    Exposure time : 2 seconds

    This image is courtesy of an Abreview submitted by Sonia Rocha

    See Abreview

  • All lanes : Anti-Cdk2 antibody [E304] (ab32147) at 1/1000 dilution (unpurified)

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate (ab27252) at 10 µg
    Lane 2 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate (ab27252) at 20 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 34 kDa
    Observed band size : 34 kDa


    Exposure time : 4 minutes

    This blot was produced using a 10% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 5% Bovine Serum Albumin before being incubated with ab32147 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution.

Anti-Cdk2 antibody [E304] (ab32147)参考文献

This product has been referenced in:
  • Tian LQ  et al. MicroRNA-197 inhibits cell proliferation by targeting GAB2 in glioblastoma. Mol Med Rep N/A:N/A (2016). Read more (PubMed: 27035789) »
  • Hu R  et al. TMEM45B, up-regulated in human lung cancer, enhances tumorigenicity of lung cancer cells. Tumour Biol 37:12181-12191 (2016). Human . Read more (PubMed: 27225290) »

See all 7 Publications for this product

Product Wall

Application Western blot
Loading amount 20 µg
Gel Running Conditions Reduced Denaturing (10%)
Sample Human Cell lysate - whole cell (osteosarcoma cells)
Specification osteosarcoma cells
Treatment lane 1 control, lane 2 siRNA for CDK2, 48h
Blocking step Milk as blocking agent for 10 minute(s) · Concentration: 10% · Temperature: 21°C
Username

Dr. Sonia Rocha

Verified customer

提交于 Sep 26 2014

Application Immunocytochemistry/ Immunofluorescence
Blocking step Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 22°C
Sample Human Cell (MCF-7)
Specification MCF-7
Permeabilization Yes - Triton X-100
Fixative Paraformaldehyde
Username

Abcam user community

Verified customer

提交于 Aug 04 2014

Application Western blot
Loading amount 50000 cells
Gel Running Conditions Reduced Denaturing (10% gel)
Sample Human Cell lysate - whole cell (HEK293, MCF-7, MDA-MB-231)
Specification HEK293, MCF-7, MDA-MB-231
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Abcam user community

Verified customer

提交于 Aug 04 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - nuclear (T-47D)
Loading amount 60 µg
Specification T-47D
Treatment 0.08% DMSO for 48 hrs
Gel Running Conditions Non-reduced Non-Denaturing (Native) (4-20% Tris-glycine)
Blocking step (agent) for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
Username

Abcam user community

Verified customer

提交于 Aug 31 2009

Thank you for your patience. Regarding ab32147 (Cdk2 antibody), the exact immunogen sequence and amino acid location is proprietary information. At this time, we have not tested ab32147 for cross-reactivity with Cdk1. Please don't hesitate to co...

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"