重组Anti-ATG7抗体[EP1759Y] (ab52472)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EP1759Y] to ATG7
- Suitable for: Flow Cyt (Intra), ICC/IF, WB, IP, IHC-P
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
概述
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产品名称
Anti-ATG7抗体[EP1759Y]
参阅全部 ATG7 一抗 -
描述
兔单克隆抗体[EP1759Y] to ATG7 -
宿主
Rabbit -
经测试应用
适用于: Flow Cyt (Intra), ICC/IF, WB, IP, IHC-Pmore details -
种属反应性
与反应: Human -
免疫原
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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阳性对照
- IHC-P: Human cervical carcinomaWB: Jurkat, HepG2, HEK293 and HAP1 cell lysate ICC/IF: HT-29 and HeLa whole cell lysate (ab150035)Flow Cyt (intra): HEK293 and HeLa 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.
We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.
Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.
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. Avoid freeze / thaw cycle. -
存储溶液
pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.21% BSA -
Concentration information loading...
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纯度
Protein A purified -
克隆
单克隆 -
克隆编号
EP1759Y -
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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Positive Controls
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab52472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) |
1/50 - 1/100.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
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ICC/IF |
1/100 - 1/500.
For unpurified use at 10 µg/mL. |
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WB | (2) |
1/100000 - 1/200000. Detects a band of approximately 70 kDa (predicted molecular weight: 78 kDa).
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IP |
1/30 - 1/50.
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IHC-P |
1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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说明 |
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Flow Cyt (Intra)
1/50 - 1/100. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
ICC/IF
1/100 - 1/500. For unpurified use at 10 µg/mL. |
WB
1/100000 - 1/200000. Detects a band of approximately 70 kDa (predicted molecular weight: 78 kDa). |
IP
1/30 - 1/50. |
IHC-P
1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
靶标
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功能
E1-like activating enzyme involved in the 2 ubiquitin-like systems required for cytoplasm to vacuole transport (Cvt) and autophagy. Activates ATG12 for its conjugation with ATG5 as well as the ATG8 family proteins for their conjugation with phosphatidylethanolamine. Both systems are needed for the ATG8 association to Cvt vesicles and autophagosomes membranes. Required for autophagic death induced by caspase-8 inhibition. Required for mitophagy which contributes to regulate mitochondrial quantity and quality by eliminating the mitochondria to a basal level to fulfill cellular energy requirements and preventing excess ROS production. Modulates p53/TP53 activity to regulate cell cycle and survival during metabolic stress. Plays also a key role in the maintenance of axonal homeostasis, the prevention of axonal degeneration, the maintenance of hematopoietic stem cells, the formation of Paneth cell granules, as well as in adipose differentiation. -
组织特异性
Widely expressed, especially in kidney, liver, lymph nodes and bone marrow. -
序列相似性
Belongs to the ATG7 family. -
结构域
The C-terminal part of the protein is essential for the dimerization and interaction with ATG3 and ATG12.
The N-terminal FAP motif (residues 15 to 17) is essential for the formation of the ATG89-PE and ATG5-ATG12 conjugates. -
翻译后修饰
Acetylated by EP300. -
细胞定位
Cytoplasm. Preautophagosomal structure. Localizes also to discrete punctae along the ciliary axoneme and to the base of the ciliary axoneme. - Information by UniProt
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数据库链接
- Entrez Gene: 10533 Human
- Omim: 608760 Human
- SwissProt: O95352 Human
- Unigene: 38032 Human
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别名
- 1810013K23Rik antibody
- Apg 7 antibody
- APG7 autophagy 7 like antibody
see all
图片
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All lanes : Anti-ATG7 antibody [EP1759Y] (ab52472) at 1/100000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : ATG7 knockout HeLa cell lysate
Lane 3 : HepG2 cell lysate
Lane 4 : Jurkat cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 78 kDa
Observed band size: 75 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-ATG7 antibody [EP1759Y] staining at 1/100000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab52472 was shown to bind specifically to ATG7. A band was observed at 75 kDa in wild-type HeLa cell lysates with no signal observed at this size in ATG7 knockout cell line ab283307 (knockout cell lysate ab287353). To generate this image, wild-type and ATG7 knockout HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
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Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: ATG7 knockout HAP1 cell lysate (20 µg)
Lane 3: Jurkat cell lysate (20 µg)
Lane 4: HepG2 cell lysate (20 µg)Lanes 1 - 4: Merged signal (red and green). Green - ab52472 observed at 77 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab52472 was shown to specifically react with ATG7 when ATG7 knockout samples were used. Wild-type and ProteinX knockout samples were subjected to SDS-PAGE. ab52472 and ab8245 (loading control to Apg7) were both diluted 1/2000 and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging. -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATG7 antibody [EP1759Y] (ab52472)
Immunohistochemical analysis of paraffin-embedded human cervical carcinoma sections labelling ATG7 with purified ab52472 at a dilution of 1/500. The secondary antibody used was ab97051, Goat Anti-Rabbit IgG H&L (HRP) at a dilution of 1/500. The sample was counterstained with hematoxylin. Antigen retrieval was performed using EDTA Buffer; pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.
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Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling ATG7 with purified ab52472 at 1/100. Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% Triton X-100. ab150077, Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. Cells were counter-stained with ab7291 anti-Tubulin (mouse mAb) followed by ab150120, AlexaFluor®594 goat anti-mouse secondary both at 1/1000. Nuclei were counterstained with DAPI (blue).
For negative control 1, rabbit primary antibody was used followed by anti-mouse secondary antibody (ab150120). For negative control 2, ab7291 (mouse primary antibody) was used followed by anti-rabbit secondary antibody (ab150077).
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Intracellular Flow Cytometry analysis of HeLa cells labelling ATG7 (red) with purified ab52472 at dilution of 1/100. The secondary antibody used was goat anti rabbit IgG (FITC) at 1/500. Cells were fixed with 4% paraformaldehyde. Isotype control antibody was Rabbit monoclonal IgG (black). The blue line shows cells without incubation with primary antibody and secondary antibody.
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ab52472 at 1/30 dilution immunoprecipitating ATG7 in HEK293 whole cell lysate observed at 70 KDa (lanes 1 and 2).
Lane 1 (input): HEK293 whole cell lysate 10ug
Lane 2 (+): ab52472 + HEK293 whole cell lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab52472 in HEK293 whole cell lysate
For western blotting, ab52472 was used followed by VeriBlot for IP Detection Reagent (HRP) (ab131366) for detection at a dilution of 1/10,000.
Blocking and Diluting buffer and concentration: 5% NFDM/TBST.
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All lanes : Anti-ATG7 antibody [EP1759Y] (ab52472) at 1/100000 dilution
Lane 1 : HEK293 whole cell lysate
Lane 2 : HepG2 whole cell lysate
Lane 3 : Jurkat whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 78 kDa
Observed band size: 70 kDa why is the actual band size different from the predicted?
Blocking and Diluting buffer 5% NFDM/TBST -
Immunocytochemistry/Immunofluorescence analysis of HT-29 (human colorectal adenocarcinoma) cells labelling ATG7 with purified ab52472 at 1/500. Cells were fixed with 100% methanol. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG (ab150077) at 1/1000 dilution was used as the secondary antibody. Nuclei couterstained with DAPI (blue).
Secondary Only Control: PBS was used instead of the primary antibody as the negative control.
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Overlay histogram showing HEK293 cells stained with unpurified ab52472 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab52472, 1/50 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HEK293 cells fixed with 100% methanol (5 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.
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
文献 (57)
ab52472 被引用在 57 文献中.
- Wu MD et al. Acetylshikonin induces autophagy-dependent apoptosis through the key LKB1-AMPK and PI3K/Akt-regulated mTOR signalling pathways in HL-60 cells. J Cell Mol Med 26:1606-1620 (2022). PubMed: 35106915
- Chen H et al. Exposure of zebrafish to a cold environment triggered cellular autophagy in zebrafish liver. J Fish Dis 45:991-1000 (2022). PubMed: 35395109
- Huang H et al. L3MBTL2-mediated CGA transcriptional suppression promotes pancreatic cancer progression through modulating autophagy. iScience 25:104249 (2022). PubMed: 35521536
- Wang C et al. Long non-coding RNAs LINC00689 inhibits the apoptosis of human nucleus pulposus cells via miR-3127-5p/ATG7 axis-mediated autophagy. Open Med (Wars) 17:1821-1832 (2022). PubMed: 36475062
- Li Y et al. The lncRNA HOTAIR regulates autophagy and affects lipopolysaccharide-induced acute lung injury through the miR-17-5p/ATG2/ATG7/ATG16 axis. J Cell Mol Med 25:8062-8073 (2021). PubMed: 34180119