Product Pathways - NF-kB Signaling
NF-κB p65 (L8F6) Mouse mAb #6956
|W IP IHC-P IF-IC F ChIP||H M R Hm Mk Mi B Dg Pg||Endogenous||65||Mouse IgG2b|
Reactivity Key: H=Human M=Mouse R=Rat Hm=Hamster Mk=Monkey Mi=Mink B=Bovine Dg=Dog Pg=Pig
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.
Specificity / Sensitivity
NF-κB p65 (L8F6) Mouse mAb recognizes endogenous levels of total NF-κB p65 protein.
Source / Purification
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human NF-κB protein.
Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-) or SignalSilence® NF-κB p65 siRNA I #6261 (+), using NF-κB p65 (L8F6) Mouse mAb (upper) or α-Tubulin (11Η10) Rabbit mAb #2125 (lower). The NF-κB p65 (L8F6) Mouse mAb confirms silencing of NF-κB p65 expression, while the α-Tubulin (11Η10) Rabbit mAb is used as a loading control.
Western blot analysis of extracts from various cell lines using NF-κB p65 (L8F6) Mouse mAb.
Immunohistochemical analysis of human chronic cholecystitis tissue using NF-κB p65 (L8F6) Mouse mAb.
Immunohistochemical analysis of paraffin-embedded OVCAR8 cell pellets treated with Human Tumor Necrosis Factor-α (hTNF-α) #8902 (left) or treated with SignalSilence® NF-κB p65 siRNA I #6261 (right), using NF-κB p65 (L8F6) Mouse mAb.
Immunohistochemical analysis of paraffin-embedded HeLa cell pellets, untreated (left) or treated with Human Tumor Necrosis Factor-α (hTNF-α) #8902 (right), using NF-κB p65 (L8F6) Mouse mAb.
Flow cytometric analysis of HeLa cells using NF-κB p65 (L8F6) Mouse mAb (blue) compared to a nonspecific negative control antibody (red).
Confocal immunofluorescent analysis of HeLa cells, untreated (left) or treated with Human Tumor Necrosis Factor-α (hTNF-α) #8902 (20 ng/mL, 20 min; right), using NF-κB p65 (L8F6) Mouse mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HeLa cells treated with Human Tumor Necrosis Factor-α (hTNF-α) #8902 (30 ng/ml, 1 hr) and either 10 μl of NF-κB p65 (L8F6) Mouse mAb or 2 μl of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human IκBα Promoter Primers #5552, human IL-8 promoter primers, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
Transcription factors of the nuclear factor κ B (NF-κB)/Rel family play a pivotal role in inflammatory and immune responses (1,2). There are five family members in mammals: RelA, c-Rel, RelB, NF-κB1 (p105/p50), and NF-κB2 (p100/p52). Both p105 and p100 are proteolytically processed by the proteasome to produce p50 and p52, respectively. Rel proteins bind p50 and p52 to form dimeric complexes that bind DNA and regulate transcription. In unstimulated cells, NF-κB is sequestered in the cytoplasm by IκB inhibitory proteins (3-5). NF-κB-activating agents can induce the phosphorylation of IκB proteins, targeting them for rapid degradation through the ubiquitin-proteasome pathway and releasing NF-κB to enter the nucleus where it regulates gene expression (6-8). NIK and IKKα (IKK1) regulate the phosphorylation and processing of NF-κB2 (p100) to produce p52, which translocates to the nucleus (9-11).
- Baeuerle, P.A. and Henkel, T. (1994) Annu Rev Immunol 12, 141-79.
- Baeuerle, P.A. and Baltimore, D. (1996) Cell 87, 13-20.
- Haskill, S. et al. (1991) Cell 65, 1281-9.
- Thompson, J.E. et al. (1995) Cell 80, 573-82.
- Whiteside, S.T. et al. (1997) EMBO J 16, 1413-26.
- Traenckner, E.B. et al. (1995) EMBO J 14, 2876-83.
- Scherer, D.C. et al. (1995) Proc Natl Acad Sci USA 92, 11259-63.
- Chen, Z.J. et al. (1996) Cell 84, 853-62.
- Senftleben, U. et al. (2001) Science 293, 1495-9.
- Coope, H.J. et al. (2002) EMBO J 21, 5375-85.
- Xiao, G. et al. (2001) Mol Cell 7, 401-9.
Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!
- 3033 Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb
- 7072 Phototope®-HRP Western Blot Detection System, Anti-mouse IgG, HRP-linked Antibody
- 7076 Anti-mouse IgG, HRP-linked Antibody
- 7720 Prestained Protein Marker, Broad Range (Premixed Format)
- 7727 Biotinylated Protein Ladder Detection Pack
- 8112 SignalStain® Antibody Diluent
- 8125 SignalStain® Boost IHC Detection Reagent (HRP, Mouse)
- 5425 Normal Goat Serum
- 9997 Tris Buffered Saline with Tween 20 (TBST-10X)
For Research Use Only. Not For Use In Diagnostic Procedures.