IKKe/IKK-i

NF-kB (nuclear factor kB) is sequestered in the cytoplasm by IkB family of inhibitory proteins that mask the nuclear localization signal of NF-kB thereby preventing translocation of NF-kB to the nucleus (1). External stimuli such as tumor necrosis factor or other cytokines results in phosphorylation and degradation of IkB releasing NF-kB dimers. NF-kB dimer subsequently translocates to the nucleus and activates target genes. Synthesis of IkBa is autoregulated (2). IkB proteins are phosphorylated by IkB kinase complex consisting of at least three proteins, IKK1/a, IKK2/b, and IKK3/g (3-5). Using a subtractive hybridization technique, a novel kinase, IKKi/IKKe has been isolated (6). LPS increases IKKi mRNA level in mouse macrophage cell lines. This protein has significant sequence homology with kinase domains of IKK1/a and IKK-2/b. Overexpression of wild type IKKi in cells phosphorylates Ser32 and Ser36 of IkBa. \n\nApplications: \nSuitable for the detection of IKKe expression by Western blotting. A band of approx. size 80 kD is detected. Daudi, Jurkat or 293 whole cell lysate can be used as a positive control. It has tested negative on mouse NIH 3T3 cells. Western blot analysis of 30ug of total cell lysate from Daudi cells with antibody at 2ug/ml dilution. \n\nRecommended Dilution: \nWestern blot: 2ug/ml\n\nStorage and Stability: \nMay be stored at 4 degrees C for short-term only. For long-term storage and to avoid repeated freezing and thawing, aliquot and add glycerol (40-50%). Freeze at -20 degrees C. Aliquots are stable for at least 12 months at -20 degrees 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.

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