84-1015-97　Raf1, CT 100ug　R0495-06

特徴

• Raf-1 associates with members of the 14–3–3 family of proteins, which appear to protect active Raf-1 from phosphatase action.
• Important regulatory phosphorylation events occur on tyrosines 340 and 341 and serines 259, 499 (activating), serine 43 (prevents Ras:GTP binding), and serine 621 (constitutive, required for activity).
• Once active, Raf-1 associates with plasma membranes independent of Ras and phosphorylates and activates MEK, initiating the MAP kinase signaling pathway.
• Applications:Suitable for use in Western Blot and Immunoprecipitation.
• Other applications not tested.
• Recommended Dilutions:Western Blot: 0.5-2ug/ml detects Raf-1 in RIPA lysates from 3T3/A31 cell lysates.
• 3T3/A31 cell lysate was resolved by electrophoresis, transferred to nitrocellulose and probed with R0495-06 (2ug/ml).
• Proteins were visualized using an IgG (HRP) goat anti-rabbit secondary antibody and a chemiluminescence detection system
• Immunoprecipitation: 4ug immunoprecipitated Raf-1.
• The immunocomplex was then used in a coupled phosphorylation assay to activate MEK1, which in turn activates MAP Kinase 2 that phosphorylates myelin basic protein in vitro
• Optimal dilutions to be determined by the researcher.
• Included Positive Antigen Control:F4209: Fibroblast Cell, 3T3/A31.
• Add 2.5ul of 2-mercaptoethanol/100ul of lysate and boil for 5 minutes to reduce the preparation.
• Load 20ug of reduced lysate per lane for minigels.
• Storage and Stability:May be stored at 4°C for short-term only.
• Aliquot to avoid repeated freezing and thawing.
• Store at -20°C.
• Aliquots are stable for 12 months after receipt.
• For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.