Cell Signaling Technology

Product Pathways - NF-kappaB Signaling

Phospho-IκB-ε (Ser18/22) Antibody #4924

Applications Reactivity Sensitivity MW (kDa) Source
W H M R (B) (Dg) Endogenous 45 Rabbit

Applications Key:  W=Western Blotting
Reactivity Key:  H=Human  M=Mouse  R=Rat  B=Bovine  Dg=Dog
Species enclosed in parentheses are predicted to react based on 100% sequence homology. Species cross-reactivity is determined by Western blot.

Specificity / Sensitivity

Phospho-IκB-ε (Ser18/22) Antibody detects endogenous levels of IκB-ε only when phosphorylated at serines 18 and 22. No cross-reactivity was detected with other family members at physiological conditions.

Source / Purification

Polyclonal antibodies are prepared by immunizing rabbits with a synthetic phospho-peptide (KLH-coupled) corresponding to residues surrounding serine 18/22 of human IκB-ε. Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western Blotting

Western blot analysis of extracts from 293 cells treated with TNF-alpha (20 ng/ml) for the indicated times, using Phospho-IκB-ε (Ser18/22) Antibody.

Background

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 IκB-α at Ser32 and Ser36 followed by proteasome-mediated degradation that results in the release and nuclear translocation of active NF-κB (3-7). IκB-α 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). Because phosphorylation of IκB-α at Ser32/36 is essential for release of active NF-κB, phosphorylation at this site is an excellent marker of NF-κB activation (1-3).

The regulation of IκB-β and IκB-ε is similar to that of IκB-α. However, the phosphorylation and ubiquitin-mediated degradation of these proteins occurs with much slower kinetics (9). IKK phosphorylation of IκB-β occurs at Ser19 and Ser23, while IκB-ε can be phosphorylated at Ser18 and Ser22 (10).

  1. Baeuerle, P.A. and Baltimore, D. (1988) Science 242, 540-6.
  2. Beg, A.A. and Baldwin, A.S. (1993) Genes Dev 7, 2064-70.
  3. Finco, T.S. et al. (1994) Proc Natl Acad Sci USA 91, 11884-8.
  4. Brown, K. et al. (1995) Science 267, 1485-8.
  5. Brockman, J.A. et al. (1995) Mol Cell Biol 15, 2809-18.
  6. Traenckner, E.B. et al. (1995) EMBO J 14, 2876-83.
  7. Chen, Z.J. et al. (1996) Cell 84, 853-62.
  8. Karin, M. and Ben-Neriah, Y. (2000) Annu Rev Immunol 18, 621-63.
  9. Hoffmann, A. et al. (2002) Science 298, 1241-1245.
  10. Shirane, M. et al. (1999) J. Biol. Chem. 274, 28169-28174.

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