Advanced phospho-ERK (Thr202/Tyr204) cellular kit
Simple, all-in-one kit for robust detection of Phospho-ERK.
Total ERK cellular kit HTRF®
This HTRF kit monitors the cellular ERK expression level and can be used as a normalization assay for the phospho-ERK kit.
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Ease-of-use
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High sensitivity
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Highly specific
Overview
The Total-ERK1/2 assay is ideal for ERK normalization with the phospho-ERK kits. It is compatible with the buffers from the phospho or Advanced phospho-ERK kits, so the same lysate can be used for fast and easy analysis of the total and the phosphorylated protein populations.
Benefits
- COMPATIBLE WITH MANY CELL TYPES
- SPECIFICITY
- DATA NORMALIZATION
Total-ERK assay principle
The Total-ERK assay quantifies the expression level of ERK in a cell lysate. Contrary to Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis or transfer. The Total-ERK assay uses two labeled antibodies: one coupled to a donor fluorophore, the other to an acceptor. Both antibodies are highly specific for a distinct epitope on the protein. In presence of ERK in a cell extract, the addition of these conjugates brings the donor fluorophore into close proximity with the acceptor and thereby generates a FRET signal. Its intensity is directly proportional to the concentration of the protein present in the sample, and provides a means of assessing the proteins expression under a no-wash assay format.
Total-ERK 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 Total ERK HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.
Total-ERK 1-plate assay protocol
Detection of total ERK 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.
Validation of total-ERK1/2 assay on various cell lines
Human (Hela, A431, HEK293, Jurkat), murine (NIH 3T3) and Hamster (CHO-K1) cells were grown in a T175 flask at 37°C, 5% CO2 for 2 days. After removal of cell culture medium, 3 mL of supplemented lysis buffer 3 were added and incubated for 45 min. Soluble supernatants were collected after 10 min centrifuging. ERK1/2 was detected using 16 µL of cell lysate. Cell lines from other species have not been tested, hence they must be evaluated case by case.
Total-ERK assay to control phosphorylation of ERK1/2
A431 cells (100,000 cells/well) were activated with EGF for 10 min, using the two-plate assay protocol of the Phospho-ERK1/2 and Total-ERK1/2 assays. As expected, results obtained show a dose-response increase of ERK1/2 phosphorylation upon EGF stimulation while ERK1/2 expression level remains constant.
MAPK/ERK cell signaling
The MAPK/ERK signaling cascade is activated by a wide variety of receptors involved in growth and differentiation. The core signal transduction cascade elicits regulation of numerous cellular processes including adhesion, migration, apoptosis, differentiation and metabolism. MEK1/2 catalyze the phosphorylation of ERK1/2 at Tyr204 and Thr202. In response, ERK phosphorylates hundreds of cytoplasmic and nuclear substrates. The wide complexity and diversity of MAPK signaling makes ERK a key regulator and major signaling node in biology.
ERK in GPCR signaling
ERK modulation plays a major role in GPCR signaling and is therefore an important measurable GPCR readout. GPCRs act via G proteins to regulate a wide range of cellular functions. Upon stimulation, these receptors activate effectors like adenylate cyclase and phospholipase C, which influence not only intracellular concentrations of second messengers like cAMP and Ca2+, but also mediate ERK1/2 phosphorylation. GPCRs have also been shown to mediate ERK1/2 activation in a G protein-independent but beta-arrestin dependent manner.
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In collaboration with Bayer - Scientific Presentations
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HTRF cellular assays - Scientific Presentations
In collaboration with the Faculty of health and medical sciences Denmark - Scientific Presentations
Open R&D: Sanofi Access Platform
In collaboration with Sanofi - Scientific Presentations
In collaboration with the Faculty of health and medical sciences Denmark - Scientific Presentations
HTRF: One technology, many uses in a pre-clinical laboratory setting
Benefits and considerations of HTRF - Scientific Presentations
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In collaboration with MRC - Scientific Presentations
Cell Signaling: Biomarkers, Phospho- & total-protein Assays - Flyers
HTRF cellular phospho-protein assays
Physiologically relevant results fo fast flowing research - Flyers
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Signaling in the immune system - Brochures
In collaboration with Molecular Devices - Posters
PI3K/AKT/mTor translational control pathway - Posters
mechanism of action of EGFR inhibitors assessed in the pancreatic BXPC3 cell line - 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
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One technology across all samples - Application Notes
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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
Understanding GPCRs is the key to improved DD
In collaboration with GEN - Guides
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 ERK total Kit / 64NRKPET
64NRKPET - Product Insert
STING HTRF offer to bridge innate and adaptive immunity
cGAS-STING signaling pathway from A to Z - Brochures
Product Insert ERK total Kit / 64NRKPEG-64NRKPEH
64NRKPEG-64NRKPEH - 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
An innate and adaptive immunity recap
Insight into the diversity of cells & signaling pathways - Guides
How to run a cell based phospho HTRF assay
3' video to set up your Phospho assay - Videos
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The essential guide for extending your knowledge on the molecular mechanisms of neurodegenerative diseases - Guides
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Discover this infographic design on neurodegenerative diseases - Infographics
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See published experiments and data demonstrating how immunoassays rise to the challenge of astrocyte studies in neuroinflammation research - Application Notes
On-demand webinar: Linking Neuroinflammation and Neurodegeneration
New insight into neuroinflammation research - Videos
Assays for autoimmunity research
Advance your research on autoimmune diseases - Flyers
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The knowledge and data necessary to understand the inhibition value of FGFR receptors involved in human cancers. HTRF technology enables the monitoring of the activity of the AZD4547 inhibitor on cell signal transduction in 3 cancer cell lines. - Application Notes
a convincing demonstration of the reliability of the AlphaLISA and HTRF KRAS portfolios to evaluate compound in vitro therapeutic profiles in a cellular context - Application Notes
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
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Learn how to reduce time and sample consumption - Application Notes
Safety Data Sheet (DEU) ERK total Kit / 64NRKPEG
64NRKPEG - Safety Data Sheet
Safety Data Sheet (ELL) ERK total Kit / 64NRKPEG
64NRKPEG - Safety Data Sheet
Safety Data Sheet (FRA-FR) ERK total Kit / 64NRKPEG
64NRKPEG - Safety Data Sheet
Safety Data Sheet (ITA) ERK total Kit / 64NRKPEG
64NRKPEG - Safety Data Sheet
Safety Data Sheet (SPA) ERK total Kit / 64NRKPEG
64NRKPEG - Safety Data Sheet
Safety Data Sheet (ENG-GB) ERK total Kit / 64NRKPEG
64NRKPEG - Safety Data Sheet
Safety Data Sheet (ENG-US) ERK total Kit / 64NRKPEG
64NRKPEG - Safety Data Sheet
Safety Data Sheet (DEU) ERK total Kit / 64NRKPEH
64NRKPEH - Safety Data Sheet
Safety Data Sheet (ELL) ERK total Kit / 64NRKPEH
64NRKPEH - Safety Data Sheet
Safety Data Sheet (FRA-FR) ERK total Kit / 64NRKPEH
64NRKPEH - Safety Data Sheet
Safety Data Sheet (ITA) ERK total Kit / 64NRKPEH
64NRKPEH - Safety Data Sheet
Safety Data Sheet (SPA) ERK total Kit / 64NRKPEH
64NRKPEH - Safety Data Sheet
Safety Data Sheet (ENG-GB) ERK total Kit / 64NRKPEH
64NRKPEH - Safety Data Sheet
Safety Data Sheet (ENG-US) ERK total Kit / 64NRKPEH
64NRKPEH - Safety Data Sheet
Safety Data Sheet (DEU) ERK total Kit / 64NRKPET
64NRKPET - Safety Data Sheet
Safety Data Sheet (FRA-FR) ERK total Kit / 64NRKPET
64NRKPET - Safety Data Sheet
Safety Data Sheet (ITA) ERK total Kit / 64NRKPET
64NRKPET - Safety Data Sheet
Safety Data Sheet (SPA) ERK total Kit / 64NRKPET
64NRKPET - Safety Data Sheet
Safety Data Sheet (ENG-GB) ERK total Kit / 64NRKPET
64NRKPET - Safety Data Sheet
Safety Data Sheet (ENG-US) ERK total Kit / 64NRKPET
64NRKPET - Safety Data Sheet
Safety Data Sheet (ELL) ERK total Kit / 64NRKPET
64NRKPET - Safety Data Sheet
Safety Data Sheet (DEU) ERK total Kit / 64NRKPEY
64NRKPEY - Safety Data Sheet
Safety Data Sheet (ELL) ERK total Kit / 64NRKPEY
64NRKPEY - Safety Data Sheet
Safety Data Sheet (FRA-FR) ERK total Kit / 64NRKPEY
64NRKPEY - Safety Data Sheet
Safety Data Sheet (ITA) ERK total Kit / 64NRKPEY
64NRKPEY - Safety Data Sheet
Safety Data Sheet (SPA) ERK total Kit / 64NRKPEY
64NRKPEY - Safety Data Sheet
Safety Data Sheet (ENG-GB) ERK total Kit / 64NRKPEY
64NRKPEY - Safety Data Sheet
Safety Data Sheet (ENG-US) ERK total Kit / 64NRKPEY
64NRKPEY - Safety Data Sheet
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