The PhosphoPlus® IκBα (Ser32/36) Antibody Kit provides reagents and protocols for the rapid analysis of IκBα phosphorylation at Ser32/36. The kit contains a total IκBα antibody, a phospho-specific antibody, and positive/negative control whole cell lysates, along with secondary antibodies and reagents for Western blotting.
IκBα (L35A5) Mouse mAb (Amino-terminal Antigen) #4814 detects endogenous levels of total IκBα protein. Phospho-IκBα (Ser32/36) (5A5) Mouse mAb #9246 detects endogenous levels of IκBα only when phosphorylated at Ser32/36. Neither antibody cross-reacts with other IκB family members at physiological levels.
IκBα (L35A5) Mouse mAb (Amino-terminal Antigen) #4814 is produced by immunizing mice with a GST-IκBα fusion protein corresponding the amino-terminus of human IκBα. Phospho-IκBα (Ser32/36) (5A5) Mouse mAb #9246 is produced by immunizing mice with a synthetic phosphopeptide corresponding to residues surrounding Ser32/36 of human IκBα. Total cell extracts from HeLa cells prepared without treatment serve as a negative control. Supplied in SDS Sample Buffer. Total cell extracts from HeLa cells prepared with TNF-α treatment (#2169, 20 ng/ml for 5 minutes) serve as a positive control. Supplied in SDS Sample Buffer.
The NF-κB/Rel transcription factors are present in the cytosol in an inactive state complexed with the inhibitory IκB proteins (1-3). Activation occurs via phosphorylation of IκBα at Ser32 and Ser36 followed by proteasome-mediated degradation that results in the release and nuclear translocation of active NF-κB (3-7). IκBα phosphorylation and resulting Rel-dependent transcription are activated by a highly diverse group of extracellular signals including inflammatory cytokines, growth factors, and chemokines. Kinases that phosphorylate IκB at these activating sites have been identified (8).
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc. PhosphoPlus is a trademark of Cell Signaling Technology, Inc. DRAQ5 is a registered trademark of Biostatus Limited. LumiGLO is a registered trademark of Kirkegaard & Perry Laboratories.
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