The assay is a solid-phase sandwich enzyme-linked immunosorbent assay (ELISA). A capture antibody specific for MEK1 regardless of phosphorylation status, has been coated onto the multiwell strips. Standard dilutions and samples are incubated 2 hours at RT and MEK1 antigen binds to the capture antibody. After wash, a detection antibody specific for the non-phosphorylated or phosphorylated protein is incubated 1 hr at RT, and binds to the immobilized MEL1 protein. An anti-rabbit IgG-HRP completes the four-member sandwich. The reaction is visualized by TMB substrate, followed by the stop solution. The intensity of the yellow color, measured in a multiwell plate reader at 450 nm, is directly proportional to the concentration of MEK1 in original sample. The unknown concentrations are calculated from the standard curve run with each assay
Use the multiwell plate and components incuded in the assay, following instruction provided in Technical Bulletin. Components are lot-specific and not interchangeable.
phospho-MEK1 (pSer218/pSer222) ELISA is designed to detect and quantify the level of MEK1 (pSer218/pSer222) from lysates of human and mouse cells.
MEK1, also known as ERK kinase 1, MAPK kinase 1, and MKK1, is a member of the MEK family of dual specificity protein kinases.
Features and Benefits
MEK1 ELISAs are a fast (total time 4 hours), sensitive, and specific alternative to immuniblotting and RIA. The assay is facilitated by precoated plates and ready-to-use reagents.
The sensitivity is <0.9 units/mL of MEK1 (pSer218/pSer222), which is 4 times more sensitive than immunoblotting. The specificity for dually phosphorylated MEK1 (pSer218/pSer222) was confirmed by peptide competition showing that only the dual phosphopeptide containing the phosphorylated serines could block the ELISA signal.