Flow cytometric analysis of HCT 116 cells using IKKα (3G12) Mouse mAb (blue) compared to concentration-matched Mouse (G3A1) mAb IgG1 Isotype Control #5415 (red). Anti-mouse IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4408 was used as a secondary antibody.
Western blot analysis of extracts from various cell lines using IKKβ (D30C6) Rabbit mAb.
Western blot analysis of extracts from HeLa cells, untreated or treated with TNF-α (20 ng/ml) and Calyculin A #9902 (50 nM), using Phospho-IKKγ (Ser376) Antibody #2689 (top) or IKKγ (DA10-12) Mouse mAb (bottom).
Western blot analysis of recombinant IKKε Kinase #7553 and RAW 264.7 cells, untreated or LPS-treated, using IKKε Antibody.
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
Confocal immunofluorescent analysis of HCT 116 (high expression; left) and IGROV-1 (low expression; right) cells using IKKα (3G12) Mouse mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Western blot analysis of extracts from wild-type, IKKα (-/-), and IKKβ (-/-) mouse embryonic fibroblasts (MEFs) using IKKβ (D30C6) Rabbit mAb (upper) and GAPDH (14C10) Rabbit mAb #2118 (lower).
Western blot analysis of extracts from various cell lines using IKKα (3G12) Mouse mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
Western blot analysis of HeLa cell extracts (lane 1) or IKKα knock-out HeLa cells (lane 2), using IKKα (3G12) Mouse mAb #11930 (upper), or β-actin (13E5) Rabbit mAb #4970 (lower). The absence of signal in the knockout HeLa cells confirms specificity of the antibody for IKKα.
|IKKα (3G12) Mouse mAb 11930||20 µl||
||H Mk||85||Mouse IgG1|
|IKKβ (D30C6) Rabbit mAb 8943||20 µl||
||H Mk M R||87||Rabbit IgG|
|IKKγ (DA10-12) Mouse mAb 2695||20 µl||
||H R||50||Mouse IgG1|
|IKKε Antibody 2690||20 µl||
||H M R||80||Rabbit|
|Anti-rabbit IgG, HRP-linked Antibody 7074||100 µl||
|Anti-mouse IgG, HRP-linked Antibody 7076||100 µl||
The IKK Isoform Antibody Sampler Kit provides an economical means to investigate NFkappaB signaling within the cell. The kit contains primary and secondary antibodies to perform two Western blots with each antibody.
IKKα, IKKβ, IKKγ, and IKKε antibodies detect endogenous levels of total IKKα, IKKβ, IKKγ and IKKε proteins, respectively. These antibodies do not cross-react with IKK subunits other than their specified target.
The IKKβ (D30C6) Rabbit mAb was produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human IKKβ protein. The IKKα (3G12) Mouse mAb was produced by immunizing animals with a recombinant protein specific to a fragment of human IKKα protein. The IKKγ (DA10-12) Mouse mAb was produced by immunizing animals with full length human GST-IKKγ protein. Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues at the carboxyl terminus of human IKKγ. Antibodies are purified by protein A and peptide affinity chromatography.
The NF-κB/Rel transcription factors are present in the cytosol in an inactive state, complexed with the inhibitory IκB proteins (1-3). Most agents that activate NF-κB do so through a common pathway based on phosphorylation-induced, proteasome-mediated degradation of IκB (3-7). The key regulatory step in this pathway involves activation of a high molecular weight IκB kinase (IKK) complex whose catalysis is generally carried out by three tightly associated IKK subunits. IKKα and IKKβ serve as the catalytic subunits of the kinase and IKKγ serves as the regulatory subunit (8,9). Activation of IKK depends upon phosphorylation at Ser177 and Ser181 in the activation loop of IKKβ (Ser176 and Ser180 in IKKα), which causes conformational changes, resulting in kinase activation (10-13).
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