Anti-eIF4EBP1 抗体 [Y329] (ab32024)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [Y329] to eIF4EBP1
- Suitable for: WB, IHC-P, Flow Cyt (Intra), IP, ICC/IF
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
製品の概要
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製品名
Anti-eIF4EBP1 antibody [Y329]
eIF4EBP1 一次抗体 製品一覧 -
製品の詳細
Rabbit monoclonal [Y329] to eIF4EBP1 -
由来種
Rabbit -
アプリケーション
適用あり: WB, IHC-P, Flow Cyt (Intra), IP, ICC/IFmore details -
種交差性
交差種: Mouse, Rat, Human -
免疫原
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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ポジティブ・コントロール
- WB: HeLa, HAP1, K562 and 293 cell lysates. K-562 and HEK-293 whole cell lysates. Mouse and rat skeletal muscle lysate. Rat L6 whole cell lysate. IHC-P: Human colon cancer tissue. Rat and mouse spleen tissues. Flow Cyt (intra): HAP1-WT, HEK-293, and HT1080 cells. IP: HEK-293 whole cell lysate. ICC/IF: HeLa cells.
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特記事項
Binding of eIF4EBP1 to eIF4E is reversible and is dependent on the phosphorylation status of eIF4EBP1. Non phosphorylated eIF4EBP1 will bind strongly to eIF4E while, the phosphorylated form will not. Akt, TOR, MAP kinase, S6 kinase, and Cdc2 are known kinases capable of inactivating eIF4EBP1 binding to eIF4E by phosphorylating either threonines 35, 45, 69 or serine 64. Although, not all phosphorylation events equally block the eIF4EBP1-eIF4E interaction.
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.
製品の特性
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製品の状態
Liquid -
保存方法
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. -
バッファー
pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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精製度
Protein A purified -
一次抗体 備考
Binding of eIF4EBP1 to eIF4E is reversible and is dependent on the phosphorylation status of eIF4EBP1. Non phosphorylated eIF4EBP1 will bind strongly to eIF4E while, the phosphorylated form will not. Akt, TOR, MAP kinase, S6 kinase, and Cdc2 are known kinases capable of inactivating eIF4EBP1 binding to eIF4E by phosphorylating either threonines 35, 45, 69 or serine 64. Although, not all phosphorylation events equally block the eIF4EBP1-eIF4E interaction -
ポリ/モノ
モノクローナル -
クローン名
Y329 -
アイソタイプ
IgG -
研究分野
関連製品
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Alternative Versions
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Compatible Secondaries
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Isotype control
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KO cell lines
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KO cell lysates
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Positive Controls
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Recombinant Protein
アプリケーション
The Abpromise guarantee
Abpromise保証は、 次のテスト済みアプリケーションにおけるab32024の使用に適用されます
アプリケーションノートには、推奨の開始希釈率がありますが、適切な希釈率につきましてはご検討ください。
アプリケーション | Abreviews | 特記事項 |
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WB |
1/2000. Detects a band of approximately 17 kDa (predicted molecular weight: 13 kDa).
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IHC-P |
1/100 - 1/250. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
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Flow Cyt (Intra) |
1/20.
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IP |
1/20.
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ICC/IF |
1/500.
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特記事項 |
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WB
1/2000. Detects a band of approximately 17 kDa (predicted molecular weight: 13 kDa). |
IHC-P
1/100 - 1/250. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. |
Flow Cyt (Intra)
1/20. |
IP
1/20. |
ICC/IF
1/500. |
ターゲット情報
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機能
Regulates eIF4E activity by preventing its assembly into the eIF4F complex. Mediates the regulation of protein translation by hormones, growth factors and other stimuli that signal through the MAP kinase and mTORC1 pathways. -
配列類似性
Belongs to the eIF4E-binding protein family. -
翻訳後修飾
Phosphorylated on serine and threonine residues in response to insulin, EGF and PDGF. Phosphorylation at Thr-37, Thr-46, Ser-65 and Thr-70 is regulated by mTORC1. Phosphorylated upon DNA damage, probably by ATM or ATR. - Information by UniProt
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参照データベース
- Entrez Gene: 1978 Human
- Entrez Gene: 13685 Mouse
- Entrez Gene: 116636 Rat
- Omim: 602223 Human
- SwissProt: Q13541 Human
- SwissProt: Q60876 Mouse
- SwissProt: Q62622 Rat
- Unigene: 411641 Human
see all -
別名
- 4E-BP1 antibody
- 4EBP1 antibody
- 4EBP1_HUMAN antibody
see all
画像
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All lanes : Anti-eIF4EBP1 antibody [Y329] (ab32024) at 1/2000 dilution (Purified)
Lane 1 : HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate
Lane 2 : K-562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysate
Lane 3 : Mouse skeletal muscle lysate
Lane 4 : Rat skeletal muscle lysate
Lane 5 : L6 (Rat skeletal muscle myoblast) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution
Predicted band size: 13 kDa
Observed band size: 18 kDa why is the actual band size different from the predicted?The molecular weights observed are consistent with what has been described in the literatures (PMID: 28613975, 20890458 and 27358481).
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All lanes : Anti-eIF4EBP1 antibody [Y329] (ab32024)
Lane 1 : Wild-type HAP1 cell lysate
Lane 2 : elF4EBP1 knockout HAP1 cell lysate
Lane 3 : HEK293 cell lysate
Lane 4 : K562 cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 13 kDaLanes 1 - 4: Merged signal (red and green). Green - ab32024 observed at 20 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab32024 was shown to specifically react with elF4EBP1 in wild-type Hap1 cells. Wild-type and elF4EBP1 knockout samples were subjected to SDS-PAGE. ab32024 and ab8245 (loading control to GADPH) were diluted 1/5000 and 1/2000 respectively 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. -
Purified ab32024 at 1/20 dilution (2µg) immunoprecipitating eIF4EBP1 in HEK-293 whole cell lysate.
Lane 1 (input): HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate 10µg
Lane 2 (+): ab32024 + HEK-293 whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32024 in HEK-293 whole cell lysate.
VeriBlot for IP Detection Reagent (HRP) (ab131366) (1/1000 dilution) was used for Western blotting.
Blocking Buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM/TBST.
Observed band size: 15-20 kDa -
Intracellular Flow Cytometry analysis of HEK-293 (Human embryonic kidney epithelial cell) cells labeling eIF4EBP1 with purified ab32024 at 1/20 dilution (10 µg/mL) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat spleen tissue sections labeling eIF4EBP1 with purified ab32024 at 1/200 dilution (0.53 µg/mL). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse spleen tissue sections labeling eIF4EBP1 with purified ab32024 at 1/200 dilution (0.53 µg/mL). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human colon cancer tissue sections labeling eIF4EBP1 with purified ab32024 at 1/200 dilution (0.53 µg/mL). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
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All lanes : Anti-eIF4EBP1 antibody [Y329] (ab32024) at 1/5000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : EIF4EBP1 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 13 kDa
Observed band size: 13 kDaLanes 1-2: Merged signal (red and green). Green - ab32024 observed at 13 kDa. Red - loading control ab8245 observed at 37 kDa.
ab32024 Anti-eIF4EBP1 antibody [Y329] was shown to specifically react with eIF4EBP1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264784 (knockout cell lysate ab257146) was used. Wild-type and eIF4EBP1 knockout samples were subjected to SDS-PAGE. ab32024 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 5000 Dilution and 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.
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Overlay histogram showing HAP1 wildtype (green line) and HAP1-EIF4EBP1 knockout cells (red line) stained with ab32024. The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab32024, 0.1µg/ml) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) presorbed (ab150081) at 1/2000 dilution for 30 min at 22°C. A rabbit IgG isotype control antibody (ab172730) was used at the same concentration and conditions as the primary antibody (HAP1 wildtype - black line, HAP1-EIF4EBP1 knockout - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity). Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter. This antibody can also be used in HAP1 cells fixed with 4% formaldehyde (10 min) , permeabilized with 0.1% PBS-Triton X-100 for 15 min under the same conditions.
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Immunocytochemistry/Immunofluorescence analysis of HeLa (Human epithelial cell line from cervix adenocarcinoma) labeling eIF4EBP1 with Purified ab32024 at 1/500 dilution (5 µg/ml). Cells were fixed with 4% PFA and permeabilized with 0.1% tritonX-100. ab150077 Goat anti rabbit IgG(Alexa Fluor® 488) at 1/1000 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI. PBS was used instead of the primary antibody as the negative control.
プロトコール
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.
データシートおよび資料
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SDS download
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Datasheet download
参考文献 (35)
ab32024 は 35 報の論文で使用されています。
- Zhang L & Su X Bioactive peptide inhibits acute myeloid leukemia cell proliferation by downregulating ALKBH5-mediated m6A demethylation of EIF4EBP1 and MLST8 mRNA. Cell Oncol (Dordr) 45:355-365 (2022). WB ; Human . PubMed: 35579750
- Chen J et al. mTOR inhibitor improves testosterone-induced myocardial hypertrophy in hypertensive rats. J Endocrinol 252:179-193 (2022). PubMed: 34874016
- Akusjärvi SS et al. Integrative proteo-transcriptomic and immunophenotyping signatures of HIV-1 elite control phenotype: A cross-talk between glycolysis and HIF signaling. iScience 25:103607 (2022). PubMed: 35005552
- Sawaguchi S et al. Hypomyelinating Leukodystrophy 8 (HLD8)-Associated Mutation of POLR3B Leads to Defective Oligodendroglial Morphological Differentiation Whose Effect Is Reversed by Ibuprofen. Neurol Int 14:212-244 (2022). PubMed: 35225888
- Wu Z et al. Energy deprivation-induced AMPK activation inhibits milk synthesis by targeting PrlR and PGC-1α. Cell Commun Signal 20:25 (2022). PubMed: 35248054