重组Anti-SQSTM1 / p62抗体[EPR23101-103] (ab240635)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR23101-103] to SQSTM1 / p62
- Suitable for: Flow Cyt (Intra), WB, ICC/IF, IP
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
概述
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产品名称
Anti-SQSTM1 / p62抗体[EPR23101-103]
参阅全部 SQSTM1 / p62 一抗 -
描述
兔单克隆抗体[EPR23101-103] to SQSTM1 / p62 -
宿主
Rabbit -
特异性
WB: This antibody is not suitable for using in rat tissues. We suggest loading 40µg lysate per lane in gel to obtain good signal.
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经测试应用
适用于: Flow Cyt (Intra), WB, ICC/IF, IPmore details
不适用于: IHC-P -
种属反应性
与反应: Mouse, Rat, Human -
免疫原
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
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阳性对照
- WB: U-2 OS, HeLa, MEF, NIH/3T3, PC-12, C2C12 and J774A.1 lysates. ICC/IF: HeLa, U-2 OS and MEF cells. Flow Cyt (intra): HeLa and MEF cells. IP: HeLa, U-2 OS and MEF cells.
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常规说明
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
性能
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形式
Liquid -
存放说明
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
存储溶液
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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纯度
Protein A purified -
克隆
单克隆 -
克隆编号
EPR23101-103 -
同种型
IgG -
研究领域
相关产品
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Alternative Versions
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Compatible Secondaries
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Isotype control
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Related Products
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab240635于以下的经测试应用
“应用说明”部分 下显示的仅为推荐的起始稀释度;实际最佳的稀释度/浓度应由使用者检定。
应用 | Ab评论 | 说明 |
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Flow Cyt (Intra) |
1/500.
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WB |
1/1000. Detects a band of approximately 62 kDa (predicted molecular weight: 47 kDa).
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ICC/IF |
Use a concentration of 1 µg/ml.
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IP |
1/30.
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说明 |
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Flow Cyt (Intra)
1/500. |
WB
1/1000. Detects a band of approximately 62 kDa (predicted molecular weight: 47 kDa). |
ICC/IF
Use a concentration of 1 µg/ml. |
IP
1/30. |
靶标
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功能
Adapter protein which binds ubiquitin and may regulate the activation of NFKB1 by TNF-alpha, nerve growth factor (NGF) and interleukin-1. May play a role in titin/TTN downstream signaling in muscle cells. May regulate signaling cascades through ubiquitination. Adapter that mediates the interaction between TRAF6 and CYLD (By similarity). May be involved in cell differentiation, apoptosis, immune response and regulation of K(+) channels. -
组织特异性
Ubiquitously expressed. -
疾病相关
Defects in SQSTM1 are a cause of Paget disease of bone (PDB) [MIM:602080]. PDB is a metabolic bone disease affecting the axial skeleton and characterized by focal areas of increased and disorganized bone turn-over due to activated osteoclasts. Manifestations of the disease include bone pain, deformity, pathological fractures, deafness, neurological complications and increased risk of osteosarcoma. PDB is a chronic disease affecting 2 to 3% of the population above the age of 40 years. -
序列相似性
Contains 1 OPR domain.
Contains 1 UBA domain.
Contains 1 ZZ-type zinc finger. -
结构域
The UBA domain binds specifically 'Lys-63'-linked polyubiquitin chains of polyubiquitinated substrates. Mediates the interaction with TRIM55.
The OPR domain mediates homooligomerization and interactions with PRKCZ, PRKCI, MAP2K5 and NBR1.
The ZZ-type zinc finger mediates the interaction with RIPK1. -
翻译后修饰
Phosphorylated. May be phosphorylated by PRKCZ (By similarity). Phosphorylated in vitro by TTN. -
细胞定位
Cytoplasm. Late endosome. Nucleus. Sarcomere (By similarity). In cardiac muscles localizes to the sarcomeric band (By similarity). Localizes to late endosomes. May also localize to the nucleus. Accumulates in neurofibrillary tangles and in Lewy bodies of neurons from individuals with Alzheimer and Parkinson disease respectively. Enriched in Rosenthal fibers of pilocytic astrocytoma. In liver cells, accumulates in Mallory bodies associated with alcoholic hepatitis, Wilson disease, indian childhood cirrhosis and in hyaline bodies associated with hepatocellular carcinoma. - Information by UniProt
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数据库链接
- Entrez Gene: 8878 Human
- Entrez Gene: 18412 Mouse
- Entrez Gene: 113894 Rat
- Omim: 601530 Human
- SwissProt: Q13501 Human
- SwissProt: Q64337 Mouse
- SwissProt: O08623 Rat
- Unigene: 709030 Human
see all -
别名
- A170 antibody
- DMRV antibody
- EBI 3 associated protein of 60 kDa antibody
see all
图片
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ab240635 staining SQSTM1/p62 (autophagosome) in control HeLa cells (left panel) and SQSTM1/p62 in HeLa cells treated with 100nM bafilomycin A1 (ab120497) for 18hrs (right panel). The cells were fixed with methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab240635 at 2ug/ml and ab7291 (Tubulin) at 1/1000 dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.
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ab240635 staining SQSTM1 in wild-type Hap1 cells and SQSTM1 knockout Hap1 cells treated with chloroquine (ab142116, 50μM for 24 hrs). The cells were fixed with 100% methanol (5 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab240635 at 1μg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8). -
All lanes : Anti-SQSTM1 / p62 antibody [EPR23101-103] (ab240635) at 1/1000 dilution
Lane 1 : Wild-type U-2 OS cell lysate
Lane 2 : SQSTM1 knockout U-2 OS cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 47 kDaab240635 was shown to react with SQSTM1 in wild-type U-2 OS cells in Western blot with loss of signal observed in a SQSTM1 knockout cell line. Wild-type U-2 OS and SQSTM1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab240635 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with goat anti-rabbit HRP secondary antibodies at 0.2ug/mL before imaging. These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.
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ab240635 was shown to react with SQSTM1 in wild-type U-2 OS cells in Immunocytochemistry with loss of signal observed in a SQSTM1 knockout cell line. Wild-type and knockout cells were mixed and pelleted at a 1:1 ratio on coverslips. The cells were fixed with 4% paraformaldehyde (15 min) then permeabilized with 0.1% Triton X-100 (10min) and then blocked with 1/5000. The cells were then incubated with ab240635 at 1/500 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat anti-rabbit secondary antibody to (Alexa Fluor® 555) at 0.5 μg/ml. Acquisition of the green (wild-type), red (antibody staining) and far-red (knockout) channels was performed. Representative grayscale images of the red channel are shown. Wild-type and knockout cells are outlined with yellow and magenta dashed line, respectively. Schematic representation of the mosaic strategy used is shown on the bottom-right panel. Image was acquired with a Zeiss(LSM-880). These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.
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Immunoprecipitation of SQSTM1 in U-2 OS cells. Lysates were prepared and immunoprecipitation was performed using 1.0 μg of ab240635 pre-coupled to prot.A-Sepharose beads. Samples were washed and processed for western blot with ab56416 at 1/5000. SM=10% starting material; UB=10% unbound fraction; IP=immunoprecipitate. These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.
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Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized MEF (Mouse embryo fibroblast) cells labelling SQSTM1 / p62 with ab240635 at 1/50 dilution, followed by ab150077 AlexaFluor®488 Goat anti-Rabbit secondary antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in MEF cells. ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is ab150077 AlexaFluor®488 Goat anti-Rabbit secondary at 1/1000 dilution.
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SQSTM1 / p62 was immunoprecipitated from 0.35 mg MEF (Mouse embryonic fibroblast (immortalized)) whole cell lysate with ab240635 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab240635 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/5000 dilution.
Lane 1: MEF whole cell lysate 10ug
Lane 2: ab240635 IP in MEF whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab240635 in MEF whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 3 min.
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Intracellular flow cytometric analysis of 4% paraformaldehyde fixed, 90% methanol permeabilized MEF (Mouse embryonic fibroblast (immortalized)) cells labelling SQSTM1 / p62 with ab240635 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
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All lanes : Anti-SQSTM1 / p62 antibody [EPR23101-103] (ab240635) at 1/1000 dilution
Lane 1 : HeLa (human cervix adenocarcinoma epithelial cell), whole cell lysate
Lane 2 : MEF (mouse embryonic fibroblast (immortalized)), whole cell lysate
Lane 3 : NIH/3T3 (mouse embryonic fibroblast), whole cell lysate
Lane 4 : PC-12 (rat adrenal gland pheochromocytoma), whole cell lysate
Lane 5 : C2C12 (mouse myoblasts myoblast), whole cell lysate
Lane 6 : J774A.1 (mouse reticulum cell sarcoma monocyte macrophage), whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 47 kDa
Observed band size: 62 kDa why is the actual band size different from the predicted?Blocking and dilution buffer: 5% NFDM/TBST.
Exposure times.
Lanes 1-4:3 minutes; Lanes 5-6:92 seconds.
The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 24086455).
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SQSTM1 / p62 was immunoprecipitated from 0.35 mg HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate with ab240635 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab240635 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/5000 dilution.
Lane 1: HeLa whole cell lysate 10ug
Lane 2: ab240635 IP in HeLa whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab240635 in HeLa whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 30 seconds.
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Intracellular flow cytometric analysis of 4% paraformaldehyde fixed, 90% methanol permeabilized HeLa (Human cervix adenocarcinoma epithelial cell) cells labelling SQSTM1 / p62 with ab240635 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
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Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cells labelling SQSTM1 / p62 with ab240635 at 1/50 dilution, followed by ab150077 AlexaFluor®488 Goat anti-Rabbit secondary antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in HeLa cells. ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is ab150077 AlexaFluor®488 Goat anti-Rabbit secondary at 1/1000 dilution.
实验方案
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
数据表及文件
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SDS download
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Datasheet download
Certificate of Compliance
文献 (9)
ab240635 被引用在 9 文献中.
- Jia M et al. Inhibition of PI3K/AKT/mTOR Signalling Pathway Activates Autophagy and Suppresses Peritoneal Fibrosis in the Process of Peritoneal Dialysis. Front Physiol 13:778479 (2022). PubMed: 35309056
- Zhou W et al. Exercise-induced FNDC5/irisin protects nucleus pulposus cells against senescence and apoptosis by activating autophagy. Exp Mol Med 54:1038-1048 (2022). PubMed: 35882943
- Zhang L et al. Metformin Inhibits HaCaT Cell Proliferation Under Hyperlipidemia Through Reducing Reactive Oxygen Species via FOXO3 Activation. Clin Cosmet Investig Dermatol 15:1403-1413 (2022). PubMed: 35910506
- Huang L et al. Berberine Attenuates IL-1β-Induced Damage of Nucleus Pulposus Cells via Activating the AMPK/mTOR/Ulk1 Pathway. Biomed Res Int 2022:6133629 (2022). PubMed: 35915801
- Shao Z et al. RNA-binding protein HuR suppresses senescence through Atg7 mediated autophagy activation in diabetic intervertebral disc degeneration. Cell Prolif 54:e12975 (2021). PubMed: 33372336
- Xue P et al. Deficiency of optineurin enhances osteoclast differentiation by attenuating the NRF2-mediated antioxidant response. Exp Mol Med 53:667-680 (2021). PubMed: 33864025
- Chen J et al. Inhibition of Autophagy Prevents Panax Notoginseng Saponins (PNS) Protection on Cardiac Myocytes Against Endoplasmic Reticulum (ER) Stress-Induced Mitochondrial Injury, Ca2+ Homeostasis and Associated Apoptosis. Front Pharmacol 12:620812 (2021). PubMed: 33762943
- Tao J et al. Gastrodin induces lysosomal biogenesis and autophagy to prevent the formation of foam cells via AMPK-FoxO1-TFEB signalling axis. J Cell Mol Med 25:5769-5781 (2021). PubMed: 33973365
- Wei L et al. Ginkgetin alleviates high glucose-evoked mesangial cell oxidative stress injury, inflammation, and extracellular matrix (ECM) deposition in an AMPK/mTOR-mediated autophagy axis. Chem Biol Drug Des 98:620-630 (2021). PubMed: 34148304