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Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) corresponding to Human HDAC3 (C terminal).
This product is a recombinant rabbit monoclonal antibody. 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.
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.
Our Abpromise guarantee covers the use of ab32369 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||1/5000 - 1/10000. Detects a band of approximately 49 kDa (predicted molecular weight: 49 kDa).Can be blocked with HDAC3 peptide (ab221015).|
|IHC-P||1/50. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
For unpurified use at 1/250 - 1/500.
See protocols (link: http://www.abcam.com/protocols/ihc-antigen-retrieval-protocol
|ICC/IF||1/250 - 1/500.|
|IP||1/20 - 1/50.|
Immunohistochemical analysis of paraffin-embedded human endometrial carcinoma sections labelling HDAC3 with purified ab32369 at dilution of 1:50. The secondary antibody used was ab97051; a goat anti-rabbit IgG H&L (HRP) at dilution of 1/500. The sample was counterstained with hematoxylin. Antigen retrieval was performed using EDTA Buffer; pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.
Immunocytochemistry/Immunofluorescence analysis of K562 cells labelling HDAC3 with purified ab32369 at 1/500. Cells were fixed with 4% Paraformaldehyde and permeabilized using 0.1% Triton X-100. ab150077, Alexa Fluor®488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. Cells were co-stained with ab7291, a mouse anti-tubulin antibody (1/1000) using ab150120, an Alexa Fluor®594-conjugated goat anti-mouse IgG (1/1000) as the secondary. Nuclei were counterstained with DAPI (blue).
For negative control 1, rabbit primary antibody and anti-mouse secondary antibody (ab150120) were used and for negative control 2, mouse primary antibody (ab7291) and anti-rabbit secondary antibody (ab150077) were used.
Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling HDAC3 with purified ab32369 at 1/30 dilution (10ug/mL) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.
Purified ab32369 at 1/20 immunoprecipitating HDAC3 in K562 whole cell lysate observed at 49 KDa (lanes 1 and 2).
Lane 1 (input): K562 whole cell lysate 10ug
Lane 2 (+): ab32369 + K562 whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32369 in K562 whole cell lysate
The secondary antibody used was VeriBlot for IP (HRP) (ab131366) at dilution of 1/1000.
Blocking/Diluting buffer 5% NFDM/TBST.