Anti-eNOS 抗体 [EPR23750-3] (ab252439)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR23750-3] to eNOS
- Suitable for: ICC/IF, Flow Cyt (Intra), IHC-P, mIHC, WB, IP
- Reacts with: Human
Related conjugates and formulations
製品の概要
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製品名
Anti-eNOS antibody [EPR23750-3]
eNOS 一次抗体 製品一覧 -
製品の詳細
Rabbit monoclonal [EPR23750-3] to eNOS -
由来種
Rabbit -
アプリケーション
適用あり: ICC/IF, Flow Cyt (Intra), IHC-P, mIHC, WB, IPmore details -
種交差性
交差種: Human -
免疫原
Recombinant fragment. 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: EA.hy926 and HUVEC whole cell lysates. IHC-P: Human lung carcinoma, kidney, spleen and placenta tissue. ICC/IF: HUVEC cells. Flow Cyt (intra): HeLa cells. IP: EA.hy926 whole cell lysate. mIHC: Human liver tissue.
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特記事項
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
Preservative: 0.01% Sodium azide
Constituents: 59.94% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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精製度
Protein A purified -
ポリ/モノ
モノクローナル -
クローン名
EPR23750-3 -
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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Related Products
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ab252439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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ICC/IF |
1/100.
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Flow Cyt (Intra) |
1/600.
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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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mIHC |
1/200 - 1/1000.
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WB |
1/1000. Detects a band of approximately 140 kDa (predicted molecular weight: 133 kDa).
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IP |
1/30.
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特記事項 |
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ICC/IF
1/100. |
Flow Cyt (Intra)
1/600. |
IHC-P
1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
mIHC
1/200 - 1/1000. |
WB
1/1000. Detects a band of approximately 140 kDa (predicted molecular weight: 133 kDa). |
IP
1/30. |
ターゲット情報
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機能
Produces nitric oxide (NO) which is implicated in vascular smooth muscle relaxation through a cGMP-mediated signal transduction pathway. NO mediates vascular endothelial growth factor (VEGF)-induced angiogenesis in coronary vessels and promotes blood clotting through the activation of platelets.
Isoform eNOS13C: Lacks eNOS activity, dominant-negative form that may down-regulate eNOS activity by forming heterodimers with isoform 1. -
組織特異性
Platelets, placenta, liver and kidney. -
関連疾患
Variation in NOS3 seem to be associated with susceptibility to coronary spasm. -
配列類似性
Belongs to the NOS family.
Contains 1 FAD-binding FR-type domain.
Contains 1 flavodoxin-like domain. -
翻訳後修飾
Phosphorylation by AMPK at Ser-1177 in the presence of Ca(2+)-calmodulin (CaM) activates activity. In absence of Ca(2+)-calmodulin, AMPK also phosphorylates Thr-495, resulting in inhibition of activity (By similarity). Phosphorylation of Ser-114 by CDK5 reduces activity. -
細胞内局在
Cell membrane. Membrane, caveola. Cytoplasm, cytoskeleton. Golgi apparatus. Specifically associates with actin cytoskeleton in the G2 phase of the cell cycle and which is favored by interaction with NOSIP and results in a reduced enzymatic activity. - Information by UniProt
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参照データベース
- Entrez Gene: 4846 Human
- Omim: 163729 Human
- SwissProt: P29474 Human
- Unigene: 647092 Human
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別名
- cNOS antibody
- Constitutive NOS antibody
- EC NOS antibody
see all
画像
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Fluorescence multiplex immunohistochemical analysis of human liver (formalin-fixed paraffin-embedded section). Panel A shows merged staining of anti-eNOS stained on endothelial cells (ab252439; red; Opal™570) at 1:1000 ( 1.004 μg/ml) [Panel B], anti-CD163 stained on Kupffer cells (ab213612; green; Opal™520) at 1:8000 ( 0.13 μg/ml) [Panel B], and anti-Glucose Transporter GLUT2 stained on membrane of hepatocytes (ab234440; gray; Opal™690) at 1:200 ( 3.005 μg/ml) [Panel D] on human liver. DAPI was used as a nuclear counter stain. Followed by Opal Polymer HRP Ms + Rb secondary. The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope. The section was incubated in three rounds of staining: in the order of ab234440, ab213612, and ab252439 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins was used.
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Fluorescence multiplex immunohistochemical analysis of human liver (formalin-fixed paraffin-embedded section). Panel A shows merged staining of anti-MASP2 stained on cytoplasm of hepatocytes (ab277520; gray; Opal™690) at 1:100 ( 5.22 μg/ml) [Panel B] , anti-CD163 stained on Kupffer cells (ab213612; green; Opal™520) at 1:8000 ( 0.13 μg/ml) [Panel C], and anti-eNOS tained on endothelial cells (ab252439; red; Opal™570) at 1:200 ( 3.005 μg/ml) [Panel D] on human liver. DAPI was used as a nuclear counter stain. Followed by Opal Polymer HRP Ms + Rb secondary. The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope. The section was incubated in three rounds of staining: in the order of ab277520, ab213612, and ab252439 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins was used.
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Immunohistochemical analysis of paraffin-embedded human spleen tissue labeling eNOS with ab252439 at 1/500 (1.198 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining on endothelial cells in human spleen. The section was incubated with ab252439 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HUVEC cells labelling eNOS with ab252439 at 1/100 (5.99 ug/ml) dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in HUVEC cells. Negative control: HeLa (PMID: 19559671). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - 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 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
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Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized HeLa (human cervix adenocarcinoma epithelial cell, left) /HUVEC (human umbilical vein endothelial cell, right) cells labelling eNOS with ab252439 at 1/50 dilution (1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) isotype control (Black) 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.
Negative control: Hela (PMID: 19559671).
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eNOS was immunoprecipitated from 0.35 mg EA.hy926 (human somatic cell hybrid endothelial) whole cell lysate with ab252439 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab252439 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.
Lane 1: EA.hy926 (human somatic cell hybrid endothelial) whole cell lysate 10 ug
Lane 2: ab252439 IP in EA.hy926 (human somatic cell hybrid endothelial) whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab252439 in EA.hy926 (human somatic cell hybrid endothelial) whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 6 seconds.
IP lysates were unboiled.
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Anti-eNOS antibody [EPR23750-3] (ab252439) at 1/1000 dilution + HUVEC (human umbilical vein endothelial cell) whole cell lysate at 20 µg
Secondary
Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100000 dilution
Predicted band size: 133 kDa
Observed band size: 140 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 5% NFDM/TBST.
HUVEC whole cell lysate in lane 1 is made freshly and used in WB test immediately to minimize protein degradation.
Exposure time: 6 seconds.
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Immunohistochemical analysis of paraffin-embedded human lung carcinoma tissue labeling eNOS with abab252439 at 1/500 (1.198 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining on endothelial cells in human lung carcinoma. The section was incubated with ab252439 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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All lanes : Anti-eNOS antibody [EPR23750-3] (ab252439) at 1/1000 dilution
Lane 1 : EA.hy926 (human somatic cell hybrid endothelial) whole cell lysate
Lane 2 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100000 dilution
Predicted band size: 133 kDa
Observed band size: 140 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 5% NFDM/TBST.
Negative control: HeLa (PMID:19559671).
Lysates were made freshly and used in WB test immediately to minimize protein degradation. Lysates were unboiled.
Exposure time: 6 seconds.
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Immunohistochemical analysis of paraffin-embedded human placenta tissue labeling eNOS with ab252439 at 1/500 (1.198 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining in human placenta. The section was incubated with ab252439 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunohistochemical analysis of paraffin-embedded Human kidney tissue labeling eNOS with ab252439 at 1/500 (1.198 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining on endothelial cells in human kidney. The section was incubated with ab252439 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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
Certificate of Compliance
参考文献 (0)
ab252439 は論文での使用が確認できていません。