The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 5 µg/ml.
Use a concentration of 1 µg/ml. Detects a band of approximately 105 kDa (predicted molecular weight: 105 kDa).
WWP1 is an E3 ubiquitin ligase and belongs to a family of NEDD4-like proteins. WWP1 contains 4 tandem WW domains and a HECT (homologous to the E6-associated protein carboxyl terminus) domain. WW domain-containing proteins are found in all eukaryotes and play an important role in the regulation of a wide variety of cellular functions such as protein degradation, transcription, and RNA splicing.
The HECT domain of WWP1 has been implicated in regulating the localization and stability of p53 – inhibition of WWP1 results in a decrease in p53 expression, whilst WWP1 mediated stabilization of p53 appears to be associated with an accumulation of cytoplasmic p53. WWP1 also negatively regulates the TGF beta tumor suppressor pathway by inactivating its molecular components (SMAD2, SMAD4 and TGFbetaR1). WWP1 has been implicated in both breast and prostate cancers.
WW domain containing E3 ubiquitin protein ligase 1 antibody
WW domain-containing protein 1 antibody
WWP 1 antibody
Western blot - Anti-WWP1 antibody (ab43791)
All lanes : Anti-WWP1 antibody (ab43791) at 1 µg/ml
Lane 1 : MCF7 (Human breast adenocarcinoma cell line) Whole Cell Lysate Lane 2 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
Lysates/proteins at 10 µg per lane.
Secondary Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/50000 dilution Developed using the ECL technique
Performed under reducing conditions.
Predicted band size : 105 kDa Observed band size : 105 kDa Additional bands at : 36 kDa. We are unsure as to the identity of these extra bands.
Exposure time : 8 minutes
This blot was produced using a 4-12% 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 2% Bovine Serum Albumin before being incubated with ab43791 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution ab133406.
ICC/IF image of ab43791 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab43791, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 4% formaldehyde fixed (10 min) Hek293 and HepG2 cells at 5µg/ml, and in 100% methanol fixed (5 min) HeLa, Hek293, HepG2 and MCF7 cells at 5µg/ml.
Anindya R et al. Damage-induced ubiquitylation of human RNA polymerase II by the ubiquitin ligase Nedd4, but not Cockayne syndrome proteins or BRCA1. Mol Cell28:386-97 (2007).
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