Cell Signaling Technology

Product Pathways - NF-kappaB Signaling

IκB-α (L35A5) Mouse mAb (Amino-terminal Antigen) #4814

Applications Reactivity MW (kDa) Source Isotype
W IP IHC-P IF-IC F H M R Mk B 39 Mouse IgG1

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  IF-IC=Immunofluorescence (Immunocytochemistry)  F=Flow Cytometry
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey  B=Bovine
Species enclosed in parentheses are predicted to react based on 100% sequence homology. Species cross-reactivity is determined by Western blot.

Specificity / Sensitivity

IκB-α (L35A5) Mouse mAb (Amino-terminal Antigen) detects endogenous levels of total IκB-α protein.

Source / Purification

Monoclonal antibody is produced by immunizing mice with a GST-IκB-α fusion protein corresponding the amino-terminus of human IkappaB-alpha.

Western Blotting

Western Blotting

Western blot analysis of extracts from THP-1 cells, differentiated with TPA (#9905, 80 nM for 24h) and treated with 1 μg/ml LPS for the indicated times, using Phospho-IκB-α (Ser32/36) (5A5) Mouse mAb #9246 (upper) and IκB-α (L35A5) Mouse mAb (Amino-terminal Antigen) (lower).

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa, NIH/3T3 and PC12 cells, using IkB-alpha (L35A5) Mouse mAb (Amino-terminal Antigen).

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human leiomyoma, using IkappaB-alpha (L35A5) Mouse mAb (Amino-terminal Antigen).


IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human lung carcinoma, using IkappaB-alpha (L35A5) Mouse mAb (Amino-terminal Antigen).

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human renal adenocarcinoma, using IkappaB-alpha (L35A5) Mouse mAb (Amino-terminal Antigen).

Flow Cytometry

Flow Cytometry

Flow cytometric analysis of HeLa cells, using IkappaB-alpha (L35A5) Mouse mAb (Amino-terminal Antigen) (blue) compared to a nonspecifc negative control antibody (red).


IF-IC

IF-IC

Confocal immunofluorescent analysis of HeLa cells, untreated (left), or TNF-α-treated (#2169, 10 ng/ml for 15 min, right) using IκB-α (L35A5) Mouse mAb (Amino-terminal Antigen) (red). Blue pseudocolor = DRAQ5™ (fluorescent DNA dye).

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).

  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.

Application References

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