Cell Signaling Technology

Product Pathways - NF-kB Signaling

NF-κB p65 (L8F6) Mouse mAb #6956

sc-109   sc-372   sc-8008  

No. Size Price
6956S 100 µl ( 10 western blots ) ¥3,100.00 现货查询 购买询价
6956T 20 µl ( 2 western blots ) ¥1,200.00 现货查询 购买询价
6956 carrier free & custom formulation / quantityemail request
Applications Dilution Species-Reactivity Sensitivity MW (kDa) Isotype
W 1:1000 Human,Mouse,Rat,Hamster,Monkey,Mink,Bovine,Dog,Pig, Endogenous 65 Mouse IgG2b
IP 1:100
IHC-P 1:400
F 1:200
IF-IC 1:800
ChIP 1:50

Species cross-reactivity is determined by western blot.

Applications Key: W=Western Blotting, IP=Immunoprecipitation, IHC-P=Immunohistochemistry (Paraffin), F=Flow Cytometry, IF-IC=Immunofluorescence (Immunocytochemistry), ChIP=Chromatin IP,

Specificity / Sensitivity

NF-κB p65 (L8F6) Mouse mAb recognizes endogenous levels of total NF-κB p65 protein.NF-κB p65 (L8F6) Mouse mAb 能够检测内源水平NF-κB p65/RelA 蛋白总体水平。

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.单克隆抗体是通过合成人源对应的NF-κB p65/RelA C-末端周围的肽段来免疫动物而获得。

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using NF-κB p65 (L8F6) Mouse mAb.Western免疫印迹。用NF-κB p65 (L8F6) Mouse mAb研究各种类型细胞的细胞提取液。

Chromatin IP

Chromatin IP

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.染色质免疫共沉淀。HeLa细胞培养至4 x 106,并用hTNF-α #8902 (30 ng/ml, 1 hr)处理后,将染色质交联到玻片上,然后用SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003进行免疫沉淀实验,本实验中用10 μl NF-κB p65 (L8F6) Mouse mAb 抗体或2 μl Normal Rabbit IgG #2729 。对富集的DNA做real-time PCR,所用引物为SimpleChIP® Human IκBα Promoter Primers #5552, human IL-8 promoter primers和SimpleChIP® Human α Satellite Repeat Primers #4486。每个样本中沉淀的DNA量定义为相对信号与输入的总染色质相比的数值。

IHC-P (paraffin)

IHC-P (paraffin)

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.免疫组织化学染色分析石蜡包埋HeLa细胞沉淀。未经处理(左图) 或经hTNF-α #8902 (右图)处理, 所用抗体为NF-κB p65 (L8F6) Mouse mAb。

IHC-P (paraffin)

IHC-P (paraffin)

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.免疫组织化学染色分析石蜡包埋OVCAR8细胞沉淀。经hTNF-α #8902 处理(左图) 或经SignalSilence® NF-κB p65 siRNA I #6261 (右图)处理, 所用抗体为NF-κB p65 (L8F6) Mouse mAb。

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of human chronic cholecystitis tissue using NF-κB p65 (L8F6) Mouse mAb.免疫组织化学染色分析石蜡包埋人慢性胆囊炎组织。所用抗体为NF-κB p65 (L8F6) Mouse mAb。

Western Blotting

Western Blotting

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免疫印迹。用NF-κB p65 (L8F6) Mouse mAb (上图) 或α-Tubulin (11Η10) Rabbit mAb #2125 (下图) 研究转染了100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-) 或SignalSilence® NF-κB p65 siRNA I #6261 (+)的 HeLa 细胞的细胞提取液。NF-κB p65 (L8F6) Mouse mAb 确定对NF-κB p65蛋白的沉默表达, 而 α-Tubulin (11Η10) Rabbit mAb 作为上样量的对照。

IF-IC

IF-IC

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).共聚焦免疫荧光分析HeLa细胞,未经处理 (左图)或 经hTNF-α #8902 (20 ng/ml, 20 min) (右图)处理, 所用抗体为NF-κB p65 (L8F6) Mouse mAb (绿色) 肌动蛋白微丝用DY-554 phalloidin标记(红色)。Blue pseudocolor=DRAQ5® #4084 DNA 荧光染料。

Flow Cytometry

Flow Cytometry

Flow cytometric analysis of HeLa cells using NF-κB p65 (L8F6) Mouse mAb (blue) compared to a nonspecific negative control antibody (red).流式细胞仪研究HeLa细胞,所用抗体为NF-κB p65 (L8F6) Mouse mAb (蓝色) 与非特异性的阴性对照抗体(红色)。

Background

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 is then translocated to the nucleus (9-11).转录调控因子的核因子κ B(NF-κB)/Rel 家族在炎症反应和免疫反应中发挥了至关重要的作用(1,2)。在哺乳动物中一共有5个家族 : RelA, c-Rel, RelB, NF-κB1 (p105/p50), 和 NF-κB2 (p100/p52)。 p105 和 p100 在蛋白水解酶的作用下分别形成p50 和 p52。p50 和 p52形成二聚体并结合Rel蛋白,此复合体能够结合到DNA上调控转录。在未刺激的状态下, NF-κB 在IκB抑制剂作用下在细胞质中处于非活性状态(3-5)。 NF-κB激活因子能够诱导 IκB 蛋白的磷酸化, 这就使IκB能够快速的经过泛素化-蛋白酶通路降解从而释放 NF-κB,激活的NF-κB入核调控基因的表达(6-8)。 NIK 和 IKKα (IKK1) 调节磷酸化并促使NF-κB2 (p100) 生成p52, p52入核(9-11)。

  1. Baeuerle, P.A. and Henkel, T. (1994) Annu Rev Immunol 12, 141-79.
  2. Baeuerle, P.A. and Baltimore, D. (1996) Cell 87, 13-20.
  3. Haskill, S. et al. (1991) Cell 65, 1281-9.
  4. Thompson, J.E. et al. (1995) Cell 80, 573-82.
  5. Whiteside, S.T. et al. (1997) EMBO J 16, 1413-26.
  6. Traenckner, E.B. et al. (1995) EMBO J 14, 2876-83.
  7. Scherer, D.C. et al. (1995) Proc Natl Acad Sci USA 92, 11259-63.
  8. Chen, Z.J. et al. (1996) Cell 84, 853-62.
  9. Senftleben, U. et al. (2001) Science 293, 1495-9.
  10. Coope, H.J. et al. (2002) EMBO J 21, 5375-85.
  11. Xiao, G. et al. (2001) Mol Cell 7, 401-9.

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Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

Cell Signaling Technology® is a trademark of Cell Signaling Technology, Inc.

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