The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration.
1/100 - 1/250.
1/10000. Detects a band of approximately 55 kDa (predicted molecular weight: 58 kDa).
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
FunctionReceptor-regulated SMAD (R-SMAD) that is an intracellular signal transducer and transcriptional modulator activated by TGF-beta (transforming growth factor) and activin type 1 receptor kinases. Binds the TRE element in the promoter region of many genes that are regulated by TGF-beta and, on formation of the SMAD2/SMAD4 complex, activates transcription. May act as a tumor suppressor in colorectal carcinoma.
Tissue specificityExpressed at high levels in skeletal muscle, heart and placenta.
Sequence similaritiesBelongs to the dwarfin/SMAD family. Contains 1 MH1 (MAD homology 1) domain. Contains 1 MH2 (MAD homology 2) domain.
Post-translational modificationsPhosphorylated on one or several of Thr-220, Ser-245, Ser-250, and Ser-255. In response to TGF-beta, phosphorylated on Ser-465/467 by TGF-beta and activin type 1 receptor kinases. Able to interact with SMURF2 when phosphorylated on Ser-465/467, recruiting other proteins, such as SNON, for degradation. In response to decorin, the naturally occurring inhibitor of TGF-beta signaling, phosphorylated on Ser-240 by CaMK2. Phosphorylated by MAPK3 upon EGF stimulation; which increases transcriptional activity and stability, and is blocked by calmodulin. In response to TGF-beta, ubiquitinated by NEDD4L; which promotes its degradation. Acetylated on Lys-19 by coactivators in response to TGF-beta signaling, which increases transcriptional activity. Isoform short: Acetylation increases DNA binding activity in vitro and enhances its association with target promoters in vivo. Acetylation in the nucleus by EP300 is enhanced by TGF-beta.
Cellular localizationCytoplasm. Nucleus. Cytoplasmic and nuclear in the absence of TGF-beta. On TGF-beta stimulation, migrates to the nucleus when complexed with SMAD4. On dephosphorylation by phosphatase PPM1A, released from the SMAD2/SMAD4 complex, and exported out of the nucleus by interaction with RANBP1.
Immunofluorescence staining of HeLa cells with purified ab40855 at a working dilution of 1/500, counter-stained with DAPI. The secondary antibody was an Alexa Fluor® 488 conjugated goat anti-rabbit (ab150077), used at a dilution of 1/1000. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative control is shown in bottom right hand panel - for the negative control, PBS was used instead of the primary antibody.
Overlay histogram showing PC3 cells stained with ab40855 (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 (ab40855, 1/100 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) (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.
ab40855 at a 1:100 dilution staining Smad2 in human prostate carcinoma tissue.
References for Anti-Smad2 antibody [EP784Y] (ab40855)
This product has been referenced in:
Ungefroren H et al. Rac1b negatively regulates TGF-ß1-induced cell motility in pancreatic ductal epithelial cells by suppressing Smad signalling. Oncotarget5:277-90 (2014).
Read more (PubMed: 24378395) »