Phospho-S6RP (Ser235/236) cellular kit
Highly sensitive and robust kit to detect Phospho-S6RP (Ser235/236)
HTRF Total RSK1 Cellular Kit HTRF®
The Total RSK1 kit is designed to monitor the expression level of cellular RSK1, and can be used as a normalization assay for the Phospho-RSK1 (Ser380 or T359/S363) kits.
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Ease-of-use
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Highly specific
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High sensitivity
Overview
The Total RSK1 Cellular Kit is designed for the robust quantification of Total RSK1 directly using a streamlined mix-and-read, no-wash protocol. RSK1 plays a pivotal role in cell proliferation and differentiation. This kit can be used from basic research to high throughput drug screening. Its versatility means it is suitable for many applications, ranging from basic research to the analysis of pharmacological questions in cellular models.
Benefits
- COMPATIBLE WITH MANY CELL TYPES
- SENSITIVE AND ROBUST
- SPECIFICITY
Total RSK1 Assay principle
The Total RSK1 assay measures RSK1. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The Total RSK1 assay uses 2 labeled antibodies: one with a donor fluorophore, the other with an acceptor. The first antibody was selected for its specific binding to the phosphorylated motif on the protein, and 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 protein’s phosphorylation state under a no-wash assay format.
Total RSK1 2-plate Assay Protocol
The 2 plate protocol involves culturing cells in a 96-well plate before lysis. Then lysates are transferred into a 384-well low volume detection plate before the addition of Total RSK1 HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.
Total RSK1 1-plate assay protocol
Detection of Phosphorylated c-RAF (Ser338) 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.
Total and phospho RSK1 (S380) detection in EGF-stimulated A431 cells
A431 cells were plated under 100 µl in a 96-well plates (80,000 cells/well) in complete culture medium and incubated overnight at 37°C, 5% CO2. The day after, cells were treated with 50 µl of increasing concentrations of EGF for 5 min at 37°C, 5% CO2.
After medium removal, cells were then lysed with 100 µL of supplemented lysis buffer #2 for 30 minutes at RT under gentle shaking, and 16 µL of lysate were transferred into a low volume white microplate before the addition of 2 µL of the HTRF D2 detection reagent and 2 µL HTRF Eu-K detection reagent. Total RSK1 and S380 were quantified in separate wells. The HTRF signal was recorded after ON incubation.
Total and phospho RSK1 (T359/S363) detection in EGF-stimulated A431 cells
A431 cells were plated under 100 µl in a 96-well plates (80,000 cells/well) in complete culture medium and incubated overnight at 37°C, 5% CO2. The day after, cells were treated with 50 µl of increasing concentrations of EGF for 5 min at 37°C, 5% CO2.
After medium removal, cells were then lysed with 100 µL of supplemented lysis buffer #2 for 30 minutes at RT under gentle shaking, and 16 µL of lysate were transferred into a low volume white microplate before the addition of 2 µL of the HTRF D2 detection reagent and 2 µL HTRF Eu-K detection reagent. Total RSK1 and S380 were quantified in separate wells. The HTRF signal was recorded after ON incubation.
Function and regulation of RSK1
The Ribosomal protein S6 kinase alpha-1 (RSK1) is a downstream component of the MAP kinase and ERK signaling pathway and participates to the regulation of transcription factors, cell proliferation, survival, and differentiation with Serine/Threonine kinase.
It is phosphorylated at Thr359/Ser363 by MAPK1/ERK as an initial activation, and then autophosphorylates at Ser380 which allow PDK1 binding.
Simplified pathway dissection with HTRF phospho-assays and CyBi-felix liquid handling
Analyse of PI3K/AKT/mTor translational control pathway - 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
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
HTRF assays for Oncology and Inflammation
Signaling in the Immune System - Brochures
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
TCR signaling investigation with HTRF phospho assays
Study a pathway of interest in PBMC and T cells - Application Notes
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 cRaf P-S338 Kit / 63ADK018PEG
63ADK018PEG - Product Insert
Product Insert cRaf P-S338 Kit / 63ADK018PEH
63ADK018PEH - Product Insert
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
An innate and adaptive immunity recap
Insight into the diversity of cells & signaling pathways - Guides
Product Insert RSK1 total Kit / 64RSK1TPEG-64RSK1TPEH
64RSK1TPEG-64RSK1TPEH - Product Insert
Guidelines for Cell Culture and Lysis in Different Formats Prior to HTRF Detection
Seeding and lysing recommendations for a number of cell culture vessels. - Technical Notes
Assessment of drug efficacy and toxicity by combining innovative technologies
Combination of AlphaLISA®, HTRF®, or AlphaLISA® SureFire® Ultra™ immunoassays with the ATPlite™ 1step cell viability assay - Application Notes
Methodological Aspects of Homogeneous Time-Resolved Fluorescence (HTRF)
Learn how to reduce time and sample consumption - Application Notes
Safety Data Sheet (DEU) RSK1 total Kit / 64RSK1TPEG
64RSK1TPEG - Safety Data Sheet
Safety Data Sheet (ELL) RSK1 total Kit / 64RSK1TPEG
64RSK1TPEG - Safety Data Sheet
Safety Data Sheet (FRA-FR) RSK1 total Kit / 64RSK1TPEG
64RSK1TPEG - Safety Data Sheet
Safety Data Sheet (ITA) RSK1 total Kit / 64RSK1TPEG
64RSK1TPEG - Safety Data Sheet
Safety Data Sheet (SPA) RSK1 total Kit / 64RSK1TPEG
64RSK1TPEG - Safety Data Sheet
Safety Data Sheet (ENG-GB) RSK1 total Kit / 64RSK1TPEG
64RSK1TPEG - Safety Data Sheet
Safety Data Sheet (ENG-US) RSK1 total Kit / 64RSK1TPEG
64RSK1TPEG - Safety Data Sheet
Safety Data Sheet (DEU) RSK1 total Kit / 64RSK1TPEH
64RSK1TPEH - Safety Data Sheet
Safety Data Sheet (ELL) RSK1 total Kit / 64RSK1TPEH
64RSK1TPEH - Safety Data Sheet
Safety Data Sheet (FRA-FR) RSK1 total Kit / 64RSK1TPEH
64RSK1TPEH - Safety Data Sheet
Safety Data Sheet (ITA) RSK1 total Kit / 64RSK1TPEH
64RSK1TPEH - Safety Data Sheet
Safety Data Sheet (SPA) RSK1 total Kit / 64RSK1TPEH
64RSK1TPEH - Safety Data Sheet
Safety Data Sheet (ENG-GB) RSK1 total Kit / 64RSK1TPEH
64RSK1TPEH - Safety Data Sheet
Safety Data Sheet (ENG-US) RSK1 total Kit / 64RSK1TPEH
64RSK1TPEH - Safety Data Sheet
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