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Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) corresponding to Human JNK1 + JNK2 + JNK3 (phospho T183+T183+T221).
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.
Our Abpromise guarantee covers the use of ab124956 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
|WB||1/1000 - 1/10000. Detects a band of approximately 46-54 kDa.|
|IP||1/10 - 1/100.|
|IHC-P||1/100 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. (Heat to 98°C, allow to cool for 10-20 minutes)|
|ICC||1/50 - 1/100.|
|ICC/IF||Use at an assay dependent concentration.|
Dot blot analysis of JNK1/2/3 (pT183 + pT183 + pT221) peptide (Lane 1) and JNK1/2/3 non-phospho peptide (Lane 2) labelling JNK1 + JNK2 + JNK3 (phospho T183+T183+T221) with ab124956 at a dilution of 1/1000. ab97051 (Peroxidase conjugated goat anti-rabbit IgG (H+L)) was used as the secondary antibody at a dilution of 1/100000.
Blocking and dilution buffer: 5% NFDM/TBST.
Exposure time: 3 minutes.
Immunocytochemistry/Immunofluorescence analysis of untreated, Anisomycin treated and Anisomycin + LP treated NIH/3T3 cells labelling JNK1 + JNK2 + JNK3 (phospho T183 + T183 + T221) with ab124956 at a dilution of 1/100 (left) and JNK1 + JNK2 + JNK3 with ab179461 at a dilution of 1/250 (right).
Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) were also used.
The image shows increased nuclear staining after Anisomycin (250ng/ml, 30min) treatment on NIH3T3 cells. The LP treatment decreased the increased nuclear staining caused by Anisomycin.
ab179461 was used as a Pan control for ab124956. The results showed cytoplasmic staining on untreated, Anisomycin and Anisomycin + LP treated NIH3T3 cells.
Overlay histogram showing HeLa cells stained with ab124956 (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 (ab124956, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was goat anti-rabbit Alexa Fluor® 488 IgG (H+L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (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.