Intended Use:For the development of sandwich ELISAs to measure ERK1 phosphorylated at T202/Y204 and ERK2 phosphorylated at T185/Y187 in cell lysates.
Test Principle:This BioAssay™ ELISA Kit contains the basic components required for the development of sandwich ELISAs to measure ERK1 dually phosphorylated at T202/Y204 and ERK2 dually phosphorylated at T185/Y187 in cell lysates. Immobilized capture antibodies specific for ERK1 and ERK2 bind both phosphorylated and unphosphorylated protein. After washing away unbound material, a biotinylated detection antibody specific for ERK1 or ERK2 dually phosphorylated at T202/Y204 or T185/Y187, respectively, is used to detect only phosphorylated protein, utilizing a standard Streptavidin-HRP format.
Specificity:This IC ELISA specifically recognizes ERK1 dually phosphorylated at T202 and Y204, and ERK2 dually phosphorylated at T185 and Y187. Specificity was demonstrated by Western blot analysis of the protein bound by the capture antibody supplied in the kit, cross-reactivity analysis, and peptide competition.
Cross-reactivity experiments were performed with this IC ELISA to further determine specificity. Unphosphorylated recombinant human (rh) JNK1 and rhp38a were assayed at 300ng/ml and did not cross-react or interfere in the assay. Unphosphorylated rhERK1 and rhERK2 were assayed at 300ng/ml and measured 2280pg/ml (0.78% cross-reactivity) and 3150pg/ml (1.15 % cross-reactivity), respectively.
Kit Components:Contains sufficient materials to run ELISAs on at least two 96 well plates
Phospho-ERK1/ERK2 Capture Antibody: 1440mg/ml x vial when reconstituted with 200ml of PBS
Phospho-ERK1/ERK2 Detection Antibody: 14.4mg/ml x vial when reconstituted with 1ml of IC diluentPhospho-ERK1/ERK2 Standard: 1230ng/ml x vial when reconstituted with 500ml of IC diluentStreptavidin (HRP): 1x1ml
Storage and Stability:Store components at 4°C. Stable for at least 6 months. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
