IkappaB alpha Control Cell Extracts,

The NF-kB/Rel transcription factors are present in the cytosol in an inactive state, complexed with the inhibitory IkB proteins (1-3). Activation occurs via phosphorylation of IkB-a at Ser32 and Ser36, resulting in the release and nuclear translocation of active NF-kB (3-7). IkB-a phosphorylation and resulting Rel-dependent transcription are activated by a highly diverse group of extracellular signals, including inflammatory cytokines, growth factors and chemokines. Phosphorylation of IkB-a at Ser32 and Ser36 has been shown to stimulate conjugation with ubiquitin followed by proteasome-mediated degradation of IkB, resulting in the release of active NF-kB. Kinases that phosphorylate IkB at these activating sites have been identified (8). Because phosphorylation of IkB-a at Ser32 is essential for release of active NF-kB, phosphorylation at this site is an excellent marker of NF-kB activation (1-3). Nonphosphorylated IkB-a Control Cell Extracts: Total cell extracts from HeLa cells prepared without treatment serve as a negative control. Applications: Western Blots: As controls, we recommend using 20ul of phosphorylated and nonphosphorylated IkB-a cell extracts. Optimal dilutions to be determined by the researcher. , Storage and Stability:, May be stored at 4°C for short-term only. For long-term storage, store at -20°C. Aliquots are stable for at least 6 months at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.