其它组分：
IKKα (3G12) Mouse mAb 【子货号：#11930，包装：40 µl，，运保温度：–20°C】
Phospho-IKKα/β (Ser176/180) (16A6) Rabbit mAb 【子货号：#2697，包装：40 µl，，运保温度：–20°C】
TRAF2 Antibody 【子货号：#4712，包装：40 µl，，运保温度：–20°C】
TRAF3 Antibody 【子货号：#4729，包装：40 µl，，运保温度：–20°C】
Phospho-NF-κB2 p100 (Ser866/870) Antibody 【子货号：#4810，包装：40 µl，，运保温度：–20°C】
NF-κB2 p100/p52 Antibody 【子货号：#4882，包装：40 µl，，运保温度：–20°C】
RelB (C1E4) Rabbit mAb 【子货号：#4922，包装：40 µl，，运保温度：–20°C】
NIK Antibody 【子货号：#4994，包装：40 µl，，运保温度：–20°C】
Anti-rabbit IgG, HRP-linked Antibody 【子货号：#7074，包装：100 µl，，运保温度：–20°C】
Anti-mouse IgG, HRP-linked Antibody 【子货号：#7076，包装：100 µl，，运保温度：–20°C】
描述：
Phospho-IKKα/β (Ser176/180) (16A6) Rabbit mAb能够检测在Ser176/180位点磷酸化的IKKα和在Ser177/181位点磷酸化的IKKβ。Phospho-NF-κB2 p100 (Ser866/870) Antibody能够检测转染水平的在Ser866/870位点磷酸化的NF-κB2 p100。TRAF2、TRAF3、IKKα、RelB 和 p100/p52 抗体分别检测各自目标蛋白的内源总体水平。NIK抗体检测转染水平的NIK而与修饰状态无关。phospho-IKKα/β (Ser176/180) 和 phospho-p100 (Ser866/870)抗体分别是通过合成人源 IKKα和NFκB2 p100 序列上某段肽段来免疫兔子而获得。TRAF2, TRAF3, NIK, IKKα, RelB 和 p100/p52抗体分别是通过合成人源对应TRAF2 C-末端周围的肽段，TRAF3 中间区域肽段，NIK 邻近Gly659位点周围肽段，IKKα N-末端周围肽段， RelB Ser424位点周围的肽段，p100/p52 N-末端周围的肽段来免疫兔子而获得。此多克隆抗体是通过protein A和多肽亲和层析纯化。此试剂盒包含试剂能够检测非经典NF-κB信号通路上主要成员的活性状态和总体水平，包括: TRAF2, TRAF3, NIK, IKKα, p100 和 RelB。
核转录因子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，同时p50与NF-κB家族的很多成员都能结合，p52趋向于主要与RelB结合。在如TNFα和LPS 等过多的刺激下诱导经典NF-κB信号通路，一个重要的特点是对经典的 IκB激酶(IKK)复合体(包括IKKα, IKKβ, IKKγ 和 ELKS)的激活，随后磷酸化抑制 IκB分子，并通过泛素化蛋白水解途径进行快速降解(3)。通过BAFF、CD40L和其它的刺激引起的非经典NF-κB信号通路，主要是建立在诱导磷酸化和蛋白酶介导的对NF-κB2 p100进行部分的水解成为p52，此过程被NF-κB诱导激酶(NIK) 和IKKα所调控,但是不被IKKβ或IKKγ所调控(4-6)。NIK在Ser176/180位点磷酸化IKKα(6) ，在Ser866/870位点磷酸化p100,然后招募IKKα到 p100，并且对IKKαN-末端和C-末端进行进一步的磷酸化(8),导致了对p100的泛素化和处理(9)。TNF受体相关因子分子 TRAF2和TRAF3能够负调控非经典通路(10, 11), 并且他们结合到不同的受体也在决定转导信号而激活规范通路或者非经典通路或者两者的过程中发挥了重要的作用(12)。TRAF3促进了NIK在静息细胞中的快速周转, 并且其激活诱导的降解是通路中一个重要的负调控节点(13)。此通路是B细胞成熟、激活，适当的构建外周淋巴组织和保护抵抗自身免疫所必需的(14)。
原厂资料：
Specificity / Sensitivity
The Phospho-IKKα/β (Ser176/180) (16A6) Rabbit mAb detects IKKα only when phosphorylated at Ser176/180 and IKKβ only when phosphorylated at Ser177/181. The Phospho-NF-κB2 p100 (Ser866/870) Antibody detects transfected levels of NF-κB2 p100 only when phosphorylated at Ser866/870. The TRAF2, TRAF3, IKKα, RelB, and p100/p52 antibodies detect endogenous levels of total protein of their respective targets. The NIK antibody detects transfected levels of NIK regardless of modification state.
Source / Purification
Antibodies to phospho-IKKα/β (Ser176/180) and phospho-p100 (Ser866/870) are produced by immunizing rabbits with synthetic phosphopeptides corresponding to amino acids surrounding the indicated residues of human IKKα and NFκB2 p100, respectively. Antibodies to TRAF2, TRAF3, NIK, IKKα, RelB, and p100/p52 are produced by immunizing rabbits with synthetic peptides corresponding to amino acids at the carboxy terminus of TRAF2, in a central region of TRAF3, adjacent to Gly659 of human NIK, at the amino terminus of human IKKα, surrounding Ser424 of human RelB, and at the amino terminus of human p100/p52, respectively. Polyclonal antibodies are purified by Protein A and peptide affinity chromatography.
Description
This kit contains reagents to examine the activation state and total protein levels of key components in the noncanonical NF-κB pathway: TRAF2, TRAF3, NIK, IKKα, p100, and RelB.
Background
Transcription factors of the nuclear factor κB (NF-κB)/Rel family play a pivotal role in inflammatory and immune responses (1,2). There are five family members in mammals: RelA, RelB, c-Rel, 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. The p50 and p52 products form dimeric complexes with Rel proteins. While p50 associates with many of the NF-κB family members, p52 tends to form dimers primarily with RelB. A plethora of stimuli such TNFα and LPS induce the canonical NF-κB pathway, characterized by the activation of the classical IκB Kinase (IKK) complex (containing IKKα, IKKβ, IKKγ, and ELKS), which then phosphorylates inhibitory IκB molecules, targeting them for rapid degradation through a ubiquitin-proteasome pathway (3).
The noncanonical pathway, triggered by BAFF, CD40L, and certain other stimuli, is based on the inducible phosphorylation and proteasome-mediated partial degradation of NF-κB2 p100 to p52, a process regulated by the NF-κB Inducing Kinase (NIK) and IKKα, but not IKKβ or IKKγ (4-6). NIK phosphorylates IKKα at Ser176/180 (6) and p100 at Ser866/870, then recruits IKKα to p100 where IKKα phosphorylates additional residues in the N- and C-terminus (8), leading to the ubiquitination and processing of p100 (9). The TNF Receptor Associated Factor molecules TRAF2 and TRAF3 have been shown to be negative regulators of the noncanonical pathway (10, 11), and their differential binding to receptors may also play a role in determining whether transduced signals activate the canonical pathway, noncanonical pathway, or both (12). TRAF3 promotes the rapid turnover of NIK in resting cells, and its activation-induced degradation is a key regulatory point in the pathway (13). This pathway is required for B cell maturation and activation, proper architecture of peripheral lymphoid tissue, and safeguards against autoimmunity (14).
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