Revision 1
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
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UniProt ID:

#Q9D2Y4

Entrez-Gene Id:

74568

Product Includes Quantity Reactivity MW(kDa) Isotype
Phospho-MLKL (Ser345) (D6E3G) Rabbit mAb 37333 100 µl M 54 Rabbit IgG
MLKL (D6W1K) Rabbit mAb 37705 100 µl M 54 Rabbit IgG

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

PhosphoPlus® Duets from Cell Signaling Technology (CST) provide a means to assess protein activation status. Each Duet contains an activation-state and total protein antibody to your target of interest. These antibodies have been selected from CST's product offering based upon superior performance in specified applications.

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Background

Necroptosis, a regulated pathway for necrotic cell death, is triggered by a number of inflammatory signals including cytokines in the tumor necrosis factor (TNF) family, pathogen sensors such as toll-like receptors (TLRs), and ischemic injury (1,2). The process is negatively regulated by caspases and is initiated through a complex containing the RIP1 and RIP3 kinases, typically referred to as the necrosome. Mixed lineage kinase domain-like protein (MLKL) is a pseudokinase that was identified as a downstream target of RIP3 in the necroptosis pathway (3,4). During necroptosis RIP3 is phosphorylated at Ser227, which recruits MLKL and leads to its phosphorylation at Thr357 and Ser358 (3). Knockdown of MLKL through multiple mechanisms results in inhibition of necroptosis (3-5). While the precise mechanism for MLKL-induced necroptosis is unclear, some studies have shown that necroptosis leads to oligomerization of MLKL and translocation to the plasma membrane, where it affects membrane integrity (6-9).

  1. Christofferson, D.E. and Yuan, J. (2010) Curr Opin Cell Biol 22, 263-8.
  2. Kaczmarek, A. et al. (2013) Immunity 38, 209-23.
  3. Sun, L. et al. (2012) Cell 148, 213-27.
  4. Wang, Z. et al. (2012) Cell 148, 228-43.
  5. Wu, J. et al. (2013) Cell Res 23, 994-1006.
  6. Cai, Z. et al. (2014) Nat Cell Biol 16, 55-65.
  7. Chen, X. et al. (2014) Cell Res 24, 105-21.
  8. Wang, H. et al. (2014) Mol Cell 54, 133-46.
  9. Dondelinger, Y. et al. (2014) Cell Rep 7, 971-81.

Background References

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    PhosphoPlus is a registered trademark of Cell Signaling Technology, Inc.
    All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

    限制使用

    除非 CST 的合法授书代表以书面形式书行明确同意,否书以下条款适用于 CST、其关书方或分书商提供的书品。 任何书充本条款或与本条款不同的客书条款和条件,除非书 CST 的合法授书代表以书面形式书独接受, 否书均被拒书,并且无效。

    专品专有“专供研究使用”的专专或专似的专专声明, 且未专得美国食品和专品管理局或其他外国或国内专管机专专专任何用途的批准、准专或专可。客专不得将任何专品用于任何专断或治专目的, 或以任何不符合专专声明的方式使用专品。CST 专售或专可的专品提供专作专最专用专的客专,且专用于研专用途。将专品用于专断、专防或治专目的, 或专专售(专独或作专专成)或其他商专目的而专专专品,均需要 CST 的专独专可。客专:(a) 不得专独或与其他材料专合向任何第三方出售、专可、 出借、捐专或以其他方式专专或提供任何专品,或使用专品制造任何商专专品,(b) 不得复制、修改、逆向工程、反专专、 反专专专品或以其他方式专专专专专品的基专专专或技专,或使用专品开专任何与 CST 的专品或服专专争的专品或服专, (c) 不得更改或专除专品上的任何商专、商品名称、徽专、专利或版专声明或专专,(d) 只能根据 CST 的专品专售条款和任何适用文档使用专品, (e) 专遵守客专与专品一起使用的任何第三方专品或服专的任何专可、服专条款或专似专专

    Revision 1
    #24653

    PhosphoPlus® MLKL (Ser345) Antibody Duet

    PhosphoPlus® MLKL (Ser345) Antibody Duet: Image 1 Expand Image
    使用 Phospho-MLKL (Ser345) (D6E3G) Rabbit mAb(上图)、总 MLKL (D6W1K) Rabbit mAb (Mouse Specific) #37705(中图)或 β-Actin (D6A8) Rabbit mAb #8457(下图)对未经处理的 (-) 或经过以下组合处理方法处理的 L-929 细胞提取物进行蛋白质印迹分析:Z-VAD(20 μM,先于其他混合物 30 分钟添加;+)、Mouse Tumor Necrosis Factor-α (mTNF-α) #5178(20 ng/ml,4 小时;+)、SM-164(100 nM,4 小时;+)和 necrostatin-1(Nec-1,50 μM,4 小时;+)。
    PhosphoPlus® MLKL (Ser345) Antibody Duet: Image 2 Expand Image
    使用 MLKL (D6W1K) Rabbit mAb 对不同细胞系的提取物进行蛋白质印迹法分析。
    PhosphoPlus® MLKL (Ser345) Antibody Duet: Image 3 Expand Image
    使用 Phospho-MLKL (Ser345) (D6E3G) Rabbit mAb(绿色)对未处理的(左图)、已经 Z-VAD(20 μM,30 分钟)预处理后再用 SM-164 (100 nM) 和 Mouse Tumor Necrosis Factor-α (mTNF-α) #5178(20 ng/mL,2.5 小时;中间)处理,随后用 λ 磷酸酶处理(右图)的 L-929 细胞进行共聚焦免疫荧光分析。红色 = Propidium Iodide (PI)/RNase Staining Solution #4087(DNA 荧光染料)。
    PhosphoPlus® MLKL (Ser345) Antibody Duet: Image 4 Expand Image
    对 BaF3 细胞的 MLKL 蛋白进行免疫沉淀。泳道 1 为 10% input,泳道 2 使用 Rabbit (DA1E) mAb IgG XP® Isotype Control # 3900 进行免疫沉淀,泳道 3 为 MLKL (D6W1K) Rabbit mAb。使用 MLKL (D6W1K) Rabbit mAb 进行蛋白质印迹法分析。构象特异性二抗用于避免与 IgG 的反应性。