Peptide Competition and Phosphatase Stripping. Extracts of CHO-T cells transiently transfected with wild-type human IRS-1 and treated with 100 ng/mL TPA for 30 minutes were resolved by SDS-PAGE on a 10% Tris-glycine gel and transferred to PVDF. The membrane was blocked with a 5% BSA-TBST buffer overnight at 4°C and either left untreated (2-6) or treated with lambda phosphatase (1), then incubated with the IRS-1 [pS312] antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1-3), the non-phosphopeptide corresponding to the phosphopeptide immunogen (4), a generic phosphoserine-containing peptide (5), or the phosphopeptide immunogen (6). After washing, the membrane was incubated with goat F(ab')2 anti-rabbit IgG alkaline phosphatase (Prod # ALI4405) and signals were detected using the Tropix WesternStar™ method. The data show that only the phosphopeptide corresponding to IRS-1 [pS312] blocks the antibody signal, demonstrating the specificity of the antibody. The data also show that phosphatase stripping eliminates the signal, verifying that the antibody is phospho-specific.
|Tested species reactivity||Human|
|Published species reactivity||Human|
|Host / Isotype||Rabbit / IgG|
|Immunogen||The antiserum was produced against a chemically synthesized phosphopeptide derived from a region of human IRS-1 that contains serine 312. The sequence is conserved in mouse, rat and pig.|
|Purification||Antigen affinity chromatography|
|Storage buffer||Dulbecco's PBS, pH 7.3, with 1mg/ml BSA, 50% glycerol|
|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.
|Immunohistochemistry (Paraffin) (IHC (P))||See 1 publications below|
Insulin receptor substrates (IRS) are responsible for several insulin related activities, such as glucose homeostasis, cell growth, cell transformation, apoptosis and insulin signal transduction. Serine/threonine phosphorylation of IRS-1 has been demonstrated to be a negative regulator of insulin signaling and is responsible for its degradation, although IRS-1 degradation pathways are not well understood. IRS-1 has also been shown to be constitutively activated in cancers such as breast cancer, Wilm and quote;s tumors, and adrenal cortical carcinomas, thus making IRS-1 phosphorylation and subsequent degradation an attractive therapeutic option. To date there have been four subtypes identified: IRS-1,2,3, and 4, with IRS-1 being widely expressed.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
Abnormal serine phosphorylation of insulin receptor substrate 1 is associated with tau pathology in Alzheimer's disease and tauopathies.
44-814G was used in ELISA and immunohistochemistry - paraffin section to determine the expression of abnormal serine phosphorylation of insulin receptor substrate 1 in healthy and diseased human brains
|Yarchoan M,Toledo JB,Lee EB,Arvanitakis Z,Kazi H,Han LY,Louneva N,Lee VM,Kim SF,Trojanowski JQ,Arnold SE||Acta neuropathologica (128:679)||2014|