Cell Signaling Technology

Product Pathways - NF-kB Signaling

NF-κB p65 (D14E12) XP® Rabbit mAb #8242

kappa   nf kb   nf-kb   NFkappaB   NFKB   p65   sc-109   sc-372   sc-7151  

No. Size Price
8242S 100 µl ( 10 western blots ) ¥3,580.00 现货查询 购买询价
8242T 20 µl ( 2 western blots ) ¥1,400.00 现货查询 购买询价
8242 carrier free & custom formulation / quantityemail request
Applications Dilution Species-Reactivity Sensitivity MW (kDa) Isotype
W 1:1000 Human,Mouse,Rat,Hamster,Monkey,Dog, Endogenous 65 Rabbit IgG
IP 1:100
IHC-P 1:800
F 1:1600
IF-IC 1:400
ChIP 1:100

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 (D14E12) XP® Rabbit mAb recognizes endogenous levels of total NF-κB p65/RelA protein. It does not cross react with other NF-κB/Rel family members.

NF-κB p65 (D14E12) XP® Rabbit mAb 能够检测内源NF-κB p65/RelA 蛋白总体水平。此抗体不与NF-κB/Rel 家族的其它蛋白反应。

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Glu498 of human NF-κB p65/RelA protein.

此单克隆抗体是通过合成人源对应的NF-κB p65/RelA Glu498位点周围的肽段来免疫动物而获得。

IF-IC

IF-IC

Confocal immunofluorescent analysis of HT-1080 cells, untreated (left) or treated with hTNF-α #8902 (20 ng/ml, 20 min) (right), using NF-κB p65 (D14E12) XP® Rabbit 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HT-1080 细胞核,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 (D14E12) XP® Rabbit 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。

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HeLa cells treated with hTNF-α #8902 (30 ng/ml, 1 hr) and either 5 μl of NF-κB p65 (D14E12) XP® Rabbit 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5 μl of NF-κB p65 (D14E12) XP® Rabbit 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。

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using NF-κB p65 (D14E12) XP® Rabbit mAb.Western免疫印迹。用NF-κB p65 (D14E12) XP® Rabbit mAb研究各种类型细胞的细胞提取液。

Flow Cytometry

Flow Cytometry

Flow cytometric analysis of HeLa cells using NF-κB p65 (D14E12) XP® Rabbit mAb (blue) compared to concentration matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (red).流式细胞仪研究HeLa细胞,所用抗体为NF-κB p65 (D14E12) XP® Rabbit mAb (蓝色) 浓度相匹配的 Rabbit (DA1E) mAb IgG XP® 同型对照#3900 (红色)。

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human chronic cholecystitis using NF-κB p65 (D14E12) XP® Rabbit 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 (D14E12) XP® Rabbit mAb。

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded HeLa cell pellets, control (left) or treated with hTNF-α #8902 (right), using NF-κB p65 (D14E12) XP® Rabbit 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 (D14E12) XP® Rabbit mAb。

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis using NF-κB p65 (D14E12) XP® Rabbit mAb on SignalSlide® NF-κB p65 IHC Controls #12873 (paraffin-embedded HCT116 cells, untreated (left) or treated with hTNF-α #8902 (right)).

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).

核因子κ B(NF-κB)/Rel 家族的转录调控因子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个家族成员: 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。Rel与p50和p52形成二聚体,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之后会入核发挥功能(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.
  12. Kozako, T. et al. (2015) Sci Rep 5, 11345.

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For Research Use Only. Not For Use In Diagnostic Procedures.

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

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

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

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