Analysis Note

Control3T3/A31 cell lysate.
Included Positive Antigen Control: Catalog # 12-305, 3T3/A31 lysate. Add 2.5 µL of 2-mercaptoethanol/100 µL of lysate and boil for 5 minutes to reduce the preparation. Load 20 µg of reduced lysate per lane for minigels.

Application

Research CategorySignaling

Research Sub CategoryMAP Kinases

Western Blotting Analysis:
A 1:500-1:2,000 diltuoin of this lot detected Raf-1 in 3T3/A31 cell lysates.

Immunoprecipitation: 2-4 µg of a previous lot immunoprecipitated Raf-1 from 400 µg of 3T3/A31 RIPA lysate.

Anti-Raf-1 Antibody, clone AM223 is an antibody against Raf-1 for use in IP & WB.

Disclaimer

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

General description

The Raf proteins (Raf-1, A-Raf, B-Raf) are Ser/Thr kinases with homology to the PKC family, containing an N-terminal regulatory domain and a c-terminal catalytic domain. Members of the Raf family bind to activated Ras GTPase, which results in Raf translocation to the plasma membrane and activation. Activated Raf proteins phosphorylate MEKs, and are therefore the principle transducers of signals from Ras to MAP kinase. In addition to its role in mitogenesis, Raf-1 may play a role in regulation of apoptosis and cell cycle progression. Activation of Raf-1 involves phosphorylation of Ser338/339 and Tyr340/341. Activating mutations of B-Raf that disrupt its auto-inhibition loop have been implicated in a number of cancers, including melanoma and colon cancer.

Immunogen

KLH-conjugated, synthetic peptide corresponding to the C-terminus (amino acids 637-648 (CTLTTSPRLPVF)) of human Raf-1.

Epitope: C-terminus

Linkage

Replaces: 04-412

Physical form

Cultured supernatant with 0.05% sodium azide.

Quality

Routinely evaluated by Western Blot on 3T3/A31 lysates.

Specificity

Predicted to cross-react with rat based on sequence homology.

Recognizes Raf-1, Mr 74 kDa.

Storage and Stability

Stable for 1 year at -20°C from date of receipt.
Handling Recommendations: Upon receipt, and prior to removing the cap, centrifuge the vial and gently mix the solution. Aliquot into microcentrifuge tubes and store at -20°C. Avoid repeated freeze/thaw cycles, which may damage IgG and affect product performance.

Target description

74 kDa