Phospho-FGFR2 (Tyr653/654) cellular kit
Simple and robust detection kit for Phoshpo-FGFR2 (Tyr653/654)
Phospho-EIF4E (Ser209) cellular kit HTRF®
The phospho-EIF4E (Ser209) kit enables the cell-based quantification of EIF4E protein phophorylated at Ser209 as a readout of mTor pathway activation.
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Highly specific
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Ease-of-use
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High sensitivity
Overview
The Phospho-EIF4E (Ser209) cellular assay kit measures endogenous EIF4E protein phosphorylated at Ser209 directly in your cell type of interest. EIF4E enables the expression of key genes involved in cellular proliferation, angiogenesis, and cell survival. EIF4E modulation has applications in metabolism/diabetes, neurosciences and oncology research.
Benefits
- VALIDATED ON ENDOGENOUS EXPRESSION
- SPECIFICITY
Phospho-EIF4E (Ser209) assay principle
The Phospho-EIF4E (Ser209) assay measures EIF4E when phosphorylated at Ser209. Contrary to Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis or transfer. The Phospho-EIF4E (Ser209) assay uses 2 labeled antibodies: one with a donor fluorophore, the other one with an acceptor. The first antibody is selected for its specific binding to the phosphorylated motif on the protein, the second for its ability to recognize the protein independent of its phosphorylation state. Protein phosphorylation enables an immune-complex formation involving both labeled antibodies and which brings the donor fluorophore into close proximity to the acceptor, thereby generating a FRET signal. Its intensity is directly proportional to the concentration of phosphorylated protein present in the sample, and provides a means of assessing the proteins phosphorylation state under a no-wash assay format.
Phospho-EIF4E (Ser209) 2-plate assay protocol
The 2 plate protocol involves culturing cells in a 96-well plate before lysis then transferring lysates to a 384-well low volume detection plate before adding phospho-EIF4E (Ser209) HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.
Phospho-EIF4E (Ser209) 1-plate assay protocol
Detection of Phosphorylated EIF4E (Ser209) with HTRF reagents can be performed in a single plate used for culturing, stimulation and lysis. No washing steps are required. This HTS designed protocol enables miniaturization while maintaining robust HTRF quality.
Western Blot versus HTRF assay
HEK293 cells were cultured for 1 day in a T175 flask until 80% confluency was reached.. Following lysis and centrifugation, equal amounts of lysates in serial dilutions were analyzed in a side by side comparison of Western Blot and HTRF.. The HTRF assay is determined to be 8-fold more sensitive than Western Blot.
Dose-response of CGP57380 on NIH3T3 cells
Murine NIH3T3 cells (50,000 cells/well) were incubated for 3 hours with various concentrations of CGP57380 inhibitor. After a 30 minutes lysis incubation, inhibition of eIF4E phosphorylation was measured using the HTRF phospho-eIF4E (Ser209) assay with the two-plate protocol.
Dose-response of CGP57380 on stimulated HeLa cells
Human HeLa cells (100,000 cells/well) were incubated with various concentrations of CGP57380 inhibitor, either for 30 minutess or 3 hours, followed by stimulation with 10 µM Anisomycin for 30 minutess at 37°C. After a 30 minutes lysis incubation, inhibition of eIF4E phosphorylation was measured using the HTRF phospho-eIF4E (Ser209) assay with the two-plate protocol.
Simplified Pathway for EIF4E
EIF4E, eukaryotic translation initiation factor 4E, initiates eukaryotic protein synthesis by binding to the 5 end of the mRNA cap structure and directing its association with the ribosomes. EIF4E enables the expression of key genes such as cyclins, c-myc, and vascular endothelial growth factor, involved in cell proliferation, angiogenesis and cell survival. Overexpression of phosphorylated eIF4E at Ser209 is associated with tumorgenesis, metastasis, and poor overall survival in several cancers making it a novel research target.
Simplified pathway dissection with HTRF phospho-assays and CyBi-felix liquid handling
Analyse of PI3K/AKT/mTor translational control pathway - Application Notes
Automation of HTRF® elF4E Kinase Assay Using a 3D Tumoroid-Based cell Model
MultiFlo™ FX Microplate Dispenser (BioTek) - Application Notes
Cell-based kinase assays in HTS ? potential and limitations for primary and secondary screening
In collaboration with Bayer - Scientific Presentations
Open R&D: Sanofi Access Platform
In collaboration with Sanofi - Scientific Presentations
Cisbio lysis buffer compatibility
Cell Signaling: Biomarkers, Phospho- & total-protein Assays - Flyers
HTRF cellular phospho-protein assays
Physiologically relevant results fo fast flowing research - Flyers
Cell Signaling: Biomarkers, Phospho- & total-protein assays - Flyers
p-S6RP, p-p70S6K and p-EIF4E - Validation of three new HTRF® phospho-assays for the study of translational control. - Posters
PI3K/AKT/mTor translational control pathway - Posters
Universal HTRF® phospho-protein platform: from 2D, 3D, primary cells to patient derived tumor cells
Analysis of a large panel of diverse biological samples and cellular models - Posters
From 2D, 3D cell cultures to xenografts: A smart HTRF platform to maximize anticancer drug discovery
One technology across all samples - Application Notes
HTRF phospho assays reveal subtle drug induced effects in tumor-xenografts
Tumor xenograft analysis: HTRF versus Western blot - Application Notes
HTRF cell-based phospho-protein data normalization
Valuable guidelines for efficiently analyzing and interpreting results - Application Notes
HTRF phospho-total lysis buffer: a universal alternative to RIPA lysis buffers
Increased flexibility of phospho-assays - Application Notes
Multi-tissue cellular modeling and anlysis of insulin signaling - Posters
Best practices for analyzing brain samples with HTRF® phospho assays for neurosciences
Insider Tips for successful sample treatment - Technical Notes
HTRF Alpha-tubulin Housekeeping kit
Properly interpret your compound effect - Application Notes
Optimize your HTRF cell signaling assays on tissues
HTRF and WB compatible guidelines - Technical Notes
Key guidelines to successful cell signaling experiments
Mastering the art of cell signaling assays optimization - Guides
HTRF phospho-assays reveal subtle drug-induced effects
Detailed protocol and direct comparison with WB - Posters
A single technology for 2D cells, 3D cells, and xenograft models - Posters
Best practices for analyzing tumor xenografts with HTRF phospho assays
Protocol for tumor xenograft analysis with HTRF - Technical Notes
How to run a cell based phospho HTRF assay
What to expect at the bench - Videos
Unleash the potential of your phosphorylation research with HTRF
Unmatched ease of use, sensitivity and specificity assays - Videos
Product Insert EIF4E P-S209 Kit / 64EF4PEG-64EF4PEH
64EF4PEG-64EF4PEH - Product Insert
HTRF Product Catalog 2020 July update
All your HTRF assays in one document! - Catalog
A guide to Homogeneous Time Resolved Fluorescence
General principles of HTRF - Guides
How HTRF compares to Western Blot and ELISA
Get the brochure about technology comparison. - Brochures
HTRF® cell signaling platform combined with iCell® Hepatocytes
A solution for phospho-protein analysis in metabolic disorders - Posters
Unleash the potential of your phosphorylation research with HTRF
A fun video introducing you to phosphorylation assays with HTRF - Videos
How to run a cell based phospho HTRF assay
3' video to set up your Phospho assay - Videos
Safety Data Sheet (DEU) EIF4E P-S209 Kit / 64EF4PEG
64EF4PEG - Safety Data Sheet
Safety Data Sheet (ELL) EIF4E P-S209 Kit / 64EF4PEG
64EF4PEG - Safety Data Sheet
Safety Data Sheet (FRA-FR) EIF4E P-S209 Kit / 64EF4PEG
64EF4PEG - Safety Data Sheet
Safety Data Sheet (ITA) EIF4E P-S209 Kit / 64EF4PEG
64EF4PEG - Safety Data Sheet
Safety Data Sheet (SPA) EIF4E P-S209 Kit / 64EF4PEG
64EF4PEG - Safety Data Sheet
Safety Data Sheet (ENG-GB) EIF4E P-S209 Kit / 64EF4PEG
64EF4PEG - Safety Data Sheet
Safety Data Sheet (ENG-US) EIF4E P-S209 Kit / 64EF4PEG
64EF4PEG - Safety Data Sheet
Safety Data Sheet (DEU) EIF4E P-S209 Kit / 64EF4PEH
64EF4PEH - Safety Data Sheet
Safety Data Sheet (ELL) EIF4E P-S209 Kit / 64EF4PEH
64EF4PEH - Safety Data Sheet
Safety Data Sheet (FRA-FR) EIF4E P-S209 Kit / 64EF4PEH
64EF4PEH - Safety Data Sheet
Safety Data Sheet (ITA) EIF4E P-S209 Kit / 64EF4PEH
64EF4PEH - Safety Data Sheet
Safety Data Sheet (SPA) EIF4E P-S209 Kit / 64EF4PEH
64EF4PEH - Safety Data Sheet
Safety Data Sheet (ENG-GB) EIF4E P-S209 Kit / 64EF4PEH
64EF4PEH - Safety Data Sheet
Safety Data Sheet (ENG-US) EIF4E P-S209 Kit / 64EF4PEH
64EF4PEH - Safety Data Sheet
Safety Data Sheet (DEU) EIF4E P-S209 Kit / 64EF4PEY
64EF4PEY - Safety Data Sheet
Safety Data Sheet (ELL) EIF4E P-S209 Kit / 64EF4PEY
64EF4PEY - Safety Data Sheet
Safety Data Sheet (FRA-FR) EIF4E P-S209 Kit / 64EF4PEY
64EF4PEY - Safety Data Sheet
Safety Data Sheet (ITA) EIF4E P-S209 Kit / 64EF4PEY
64EF4PEY - Safety Data Sheet
Safety Data Sheet (SPA) EIF4E P-S209 Kit / 64EF4PEY
64EF4PEY - Safety Data Sheet
Safety Data Sheet (ENG-GB) EIF4E P-S209 Kit / 64EF4PEY
64EF4PEY - Safety Data Sheet
Safety Data Sheet (ENG-US) EIF4E P-S209 Kit / 64EF4PEY
64EF4PEY - Safety Data Sheet
Plate Reader Requirement
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