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HTRF Human Phospho-MEK1 (Ser298) Detection Kit, 500 Assay Points

This HTRF kit enables the cell-based quantitative detection of MEK1 phosphorylation at Ser298.

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Feature	Specification
Application	Cell Signaling
Sample Volume	16 µL

This HTRF kit enables the cell-based quantitative detection of MEK1 phosphorylation at Ser298.

View product information

Product variants

Unit Size: 500 assay points

Part #:

64MEK1S98PEG

Unit Size: 10,000 assay points

Part #:

64MEK1S98PEH

For research use only. Not for use in diagnostic procedures. All products to be used in accordance with applicable laws and regulations including without limitation, consumption, and disposal requirements under European REACH regulations (EC 1907/2006).

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Product information

Overview

The Phospho-MEK1 (Ser298) kit measures endogenous MEK1 protein phophorylated at Ser298, to help studies of the RAS-RAF-MEK cascade in the MAPK/ERK pathway. Phosphorylated by RAF, this dual specificity mitogen-activated protein kinase is responsible for the phosphorylation of ERK, one of the most important signaling nodes, and protein kinase in cell proliferation, survival, and differentiation pathways.

How it works

Phospho-MEK1 (Ser298) assay principle

The Phospho-MEK1 (Ser298) assay measures MEK1 when phosphorylated at Ser298. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The Phospho-MEK1 (Ser298) assay uses two antibodies: one labeled with a donor fluorophore, the other one with an acceptor. The first antibody was 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 protein’s phosphorylation state under a no-wash assay format.

Phospho how it works phospho mek1 ser298 cellular assay principle

Phospho-MEK1 (Ser298) 2-plate assay protocol

The 2 plate protocol involves culturing cells in a 96-well plate before lysis, then transferring lysates into a 384-well low volume detection plate before the addition of Phospho-MEK1 (Ser298) HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.

Phospho-MEK1 (Ser298) 1-plate assay protocol

Detection of Phosphorylated MEK1 (Ser298) 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.

Phospho how it works total mek 1 plate assay protocol

Assay validation

Detection of phospho MEK1 (Ser298) on Hela cells

Hela cells were cultured in a T175 flask. At 80% of confluency, the cell culture medium was removed and 3mL of complete lysis buffer (1X) were added. The lysis step was done under gentle shaking for 30 minutes. Lysate was then collected and diluted. 16µL of each dilution were transferred into a 384-well plate for the detection of phospho MEK1 (Ser298).

Specifications

Other specifications

Application	Cell Signaling
Automation Compatible	Yes
Brand	HTRF
Detection Modality	HTRF
Lysis Buffer Compatibility	Lysis Buffer 1 Lysis Buffer 2 Lysis Buffer 3
Molecular Modification	Phosphorylation
Product Group	Kit
Sample Volume	16 µL
Shipping Conditions	Shipped in Dry Ice
Target Class	Phosphoproteins
Target Species	Human
Technology	TR-FRET
Therapeutic Area	Cardiovascular Neuroscience Oncology & Inflammation
Unit Size	500 assay points