Revision 4

#88588Store at -20C

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.
Applications:

WB, IP, IHC-P, IF-IC

REACTIVITY:

H

SENSITIVITY:

Endogenous

MW (kDa):

62

Source/Isotype:

Mouse IgG1

UniProt ID:

#Q13501

Entrez-Gene Id:

8878

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:200
Immunohistochemistry (Paraffin) 1:400 - 1:1600
Immunofluorescence (Immunocytochemistry) 1:200 - 1:800

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.

For a carrier free (BSA and azide free) version of this product see product #17274.

Specificity / Sensitivity

SQSTM1/p62 (D5L7G) Mouse mAb recognizes endogenous levels of total SQSTM1/p62 protein.

Species Reactivity:

Human

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro220 of human SQSTM1 protein.

Background

Sequestosome 1 (SQSTM1, p62) is a ubiquitin binding protein involved in cell signaling, oxidative stress, and autophagy (1-4). It was first identified as a protein that binds to the SH2 domain of p56Lck (5) and independently found to interact with PKCζ (6,7). SQSTM1 was subsequently found to interact with ubiquitin, providing a scaffold for several signaling proteins and triggering degradation of proteins through the proteasome or lysosome (8). Interaction between SQSTM1 and TRAF6 leads to the K63-linked polyubiquitination of TRAF6 and subsequent activation of the NF-κB pathway (9). Protein aggregates formed by SQSTM1 can be degraded by the autophagosome (4,10,11). SQSTM1 binds autophagosomal membrane protein LC3/Atg8, bringing SQSTM1-containing protein aggregates to the autophagosome (12). Lysosomal degradation of autophagosomes leads to a decrease in SQSTM1 levels during autophagy; conversely, autophagy inhibitors stabilize SQSTM1 levels. Studies have demonstrated a link between SQSTM1 and oxidative stress. SQSTM1 interacts with KEAP1, which is a cytoplasmic inhibitor of NRF2, a key transcription factor involved in cellular responses to oxidative stress (3). Thus, accumulation of SQSTM1 can lead to an increase in NRF2 activity.

  1. Kirkin, V. et al. (2009) Mol Cell 34, 259-69.
  2. Seibenhener, M.L. et al. (2007) FEBS Lett 581, 175-9.
  3. Komatsu, M. et al. (2010) Nat Cell Biol 12, 213-23.
  4. Bjørkøy, G. et al. (2006) Autophagy 2, 138-9.
  5. Joung, I. et al. (1996) Proc Natl Acad Sci USA 93, 5991-5.
  6. Sanchez, P. et al. (1998) Mol Cell Biol 18, 3069-80.
  7. Puls, A. et al. (1997) Proc Natl Acad Sci USA 94, 6191-6.
  8. Vadlamudi, R.K. et al. (1996) J Biol Chem 271, 20235-7.
  9. Wooten, M.W. et al. (2005) J Biol Chem 280, 35625-9.
  10. Bjørkøy, G. et al. (2005) J Cell Biol 171, 603-14.
  11. Komatsu, M. et al. (2007) Cell 131, 1149-63.
  12. Pankiv, S. et al. (2007) J Biol Chem 282, 24131-45.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation IHC-P: Immunohistochemistry (Paraffin) IF-IC: Immunofluorescence (Immunocytochemistry)

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
SignalSilence is a registered trademark of Cell Signaling Technology, Inc.
SignalStain 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 4
#88588

SQSTM1/p62 (D5L7G) Mouse mAb

Western Blotting Image 1: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
使用 SQSTM1/p62 (D5L7G) Mouse mAb #88588(上图)和 β-actin (D6A8) Rabbit mAb #8457(下图)对 HeLa 细胞(泳道 1)或 SQSTM1 敲除型细胞(泳道 2)的提取物进行蛋白印迹分析。SQSTM1 敲除型 HeLa 细胞中没有信号,这证实了抗体对 SQSTM1 的特异性。
Western Blotting Image 2: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
使用 SQSTM1/p62 (D5L7G) Mouse mAb(上图)或 β-Actin (D6A8) Rabbit mAb #8457(下图),对不同细胞系的提取物进行蛋白质印迹分析。
Western Blotting Image 3: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
使用 SQSTM1/p62 (D5L7G) Mouse mAb(上图)或 β-Actin (D6A8) Rabbit mAb #8457(下图),对未经处理的 (-)或经 Earles 基础盐溶液 (EBSS;4 小时;+) 饥饿处理的 HCT 116 细胞提取物进行蛋白质印迹分析。
Western Blotting Image 4: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
使用 SQSTM1/p62 (D5L7G) Mouse mAb(上图)或 β-Actin (D6A8) Rabbit mAb #8457(下图),对未经处理的 (-)或经氯奎宁 #14774 (50 μM,过夜;+) 处理的 Hela 细胞提取物进行蛋白质印迹分析。
Western Blotting Image 5: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
使用 SQSTM1/p62 (D5L7G) Mouse mAb(上图)或 β-Actin (D6A8) Rabbit mAb #8457(下图),对经 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-)、SignalSilence® SQSTM1/p62 siRNA I #6394 (+) 或 SignalSilence® SQSTM1/p62 siRNA II #6399 (+) 转染的 HeLa 细胞提取物进行蛋白质印迹分析。
Immunoprecipitation Image 1: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
对 PANC-1 细胞提取物 SQSTM1/p62 蛋白进行免疫沉淀。泳道 1 为 10% input,泳道 2 为 Mouse (G3A1) mAb IgG1 Isotype Control #5415,而泳道 3 为 SQSTM1/p62 (D5L7G) Mouse mAb。使用 SQSTM1/p62 (D5L7G) Mouse mAb 进行蛋白质印迹。使用轻链特异性二抗,避免与重链 IgG 反应。
Immunohistochemistry Image 1: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
使用 SQSTM1/p62 (D5L7G) Mouse mAb,对石蜡包埋的人结肠癌组织进行免疫组织化学分析。
Immunohistochemistry Image 2: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
使用 SQSTM1/p62 (D5L7G) Mouse mAb,对石蜡包埋的人肺癌组织进行免疫组织化学分析。
Immunohistochemistry Image 3: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
使用 SQSTM1/p62 (D5L7G) Mouse mAb,对石蜡包埋的人子宫内膜样腺瘤组织进行免疫组织化学分析。
Immunohistochemistry Image 4: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
使用 SQSTM1/p62 (D5L7G) Mouse mAb,对石蜡包埋的 HDLM-2(左图)和 Daudi(右图)细胞沉淀物进行免疫组织化学分析。
Immunohistochemistry Image 5: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
在对照肽(左图)或抗原特异性肽(右图)存在的情况下,使用 SQSTM1/p62 (D5L7G) Mouse mAb, 对石蜡包埋的人胆囊炎组织进行免疫组织化学分析。
Immunofluorescence Image 1: SQSTM1/p62 (D5L7G) Mouse mAb Expand Image
使用 SQSTM1/p62 (D5L7G) Mouse mAb(绿色),对未经处理的(左图)或经氯奎宁处理(50 μM,过夜;右图)的 HeLa 细胞进行共聚焦免疫荧光分析。蓝色伪彩 = DRAQ5® #4084(DNA 荧光染料)。