Peptide Competition. Extracts of NIH3T3 cells unstimulated (1) or stimulated with 50 ng/mL PDGF for 5 minutes (lanes 2-6) were resolved by SDS-PAGE on a 10% Tris-glycine gel and transferred to PVDF. The membrane was then left untreated (lanes 1-5) or treated with lambda phosphatase (6), blocked with a 5% BSA-TBST buffer for one hour at room temperature, then incubated with the p70-S6K [pT389l antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1, 2, 6), the non-phosphorylated peptide corresponding to the phosphopeptide immunogen (3), a generic phosphothreonine containing peptide (4), or the phosphopeptide immunogen (5). After washing, the membrane was incubated with goat F(ab and quote;)2 anti-rabbit IgG HRP-conjugate (Cat. no. ALI4404) and signals were detected using the Pierce SuperSignal™ method. The data show the phosphorylation signal is increased upon stimulation and that only the phosphopeptide corresponding to p70-S6K [pT389l blocks the antibody signal, demonstrating the specificity of the antibody. The data also show that phosphatase stripping eliminates the signal, further verifying that the antibody is phospho-specific.
|Tested species reactivity||Human, Mouse, Rat|
|Host / Isotype||Rabbit / IgG|
|Immunogen||The antiserum was produced against a chemically synthesized phosphopeptide derived from the region of human p70-S6K that contains threonine 389. The sequence is conserved among multiple species including mouse and rat.|
|Purification||Antigen affinity chromatography|
|Storage buffer||Dulbecco's PBS, pH 7.3, with 50% glycerol, 1mg/ml BSA|
|Contains||0.05% sodium azide|
|Tested Applications||Dilution *|
|Western Blot (WB)||Assay Dependent|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
p70 Ribosomal Protein S6 Kinase (p70S6K) is a 70 kDa member of the ribosomal S6 kinase (RSK) family of serine/threonine kinases. It is predominantly localized in the cytoplasm, and is essential in growth factors regulated cell proliferation, pathways involving cell motility, such as metastases, the immune response, and tissue repair. p70S6K acts downstream of phosphoinositide (PI) 3-kinase, and its main physiological target is the S6 ribosomal protein, which is involved in upregulation of protein synthesis.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.