Recombinant Anti-eNOS antibody [EPR26335-79] (BSA and Azide free) (ab300072)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR26335-79] to eNOS - BSA and Azide free
- Suitable for: Flow Cyt (Intra), IHC-P, WB, ICC/IF
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
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Product name
Anti-eNOS antibody [EPR26335-79] (BSA and Azide free)
See all eNOS primary antibodies -
Description
Rabbit monoclonal [EPR26335-79] to eNOS - BSA and Azide free -
Host species
Rabbit -
Tested applications
Suitable for: Flow Cyt (Intra), IHC-P, WB, ICC/IFmore details
Unsuitable for: IP -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: Whole cell fresh lysates: HUVEC (human umbilical vein endothelial cell), EA.hy926 (human endothelial somatic cell ), bEnd.3 (mouse brain endothelioma). IHC-P: Tissues: Mouse and rat spleen and colon, rat stomach. ICC/IF: bEnd.3 (mouse brain endothelioma endothelialpolyoma middle T antigen transformed). Flow cyt. intr.: bEnd.3 (mouse brain endothelioma endothelialpolyoma middle T antigen transformed).
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General notes
ab300072 is a carrier free version of ab300071.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
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.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C. -
Storage buffer
pH: 7.2
Constituent: 100% PBS -
Carrier free
Yes -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR26335-79 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab300072 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
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Flow Cyt (Intra) |
Use at an assay dependent concentration.
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IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
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WB |
Use at an assay dependent concentration. Detects a band of approximately 140 kDa (predicted molecular weight: 133 kDa).
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ICC/IF |
Use at an assay dependent concentration.
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Notes |
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Flow Cyt (Intra)
Use at an assay dependent concentration. |
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. |
WB
Use at an assay dependent concentration. Detects a band of approximately 140 kDa (predicted molecular weight: 133 kDa). |
ICC/IF
Use at an assay dependent concentration. |
Target
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Function
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. -
Tissue specificity
Platelets, placenta, liver and kidney. -
Involvement in disease
Variation in NOS3 seem to be associated with susceptibility to coronary spasm. -
Sequence similarities
Belongs to the NOS family.
Contains 1 FAD-binding FR-type domain.
Contains 1 flavodoxin-like domain. -
Post-translational
modificationsPhosphorylation 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. -
Cellular localization
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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Database links
- Entrez Gene: 4846 Human
- Entrez Gene: 18127 Mouse
- Entrez Gene: 24600 Rat
- Omim: 163729 Human
- SwissProt: P29474 Human
- SwissProt: P70313 Mouse
- SwissProt: Q62600 Rat
- Unigene: 647092 Human
see all -
Alternative names
- cNOS antibody
- Constitutive NOS antibody
- EC NOS antibody
see all
Images
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All lanes : Anti-eNOS antibody [EPR26335-79] (ab300071) at 1/1000 dilution
Lane 1 : HUVEC (human umbilical vein endothelial cell) whole cell fresh lysate
Lane 2 : EA.hy926 (human endothelial somatic cell ) whole cell fresh lysate
Lane 3 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell fresh 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: 133 kDa
Observed band size: 140 kDa why is the actual band size different from the predicted?This data was developed using ab300071, the same antibody clone in a different buffer formulation.
5% NFDM/TBST was used as blocking and diluting buffer.
Exposure time:
Lane 1-2: 48 seconds
Lane 3: 3 minutesNegative control: HeLa (PMID:19559671)
Lysates were freshly made and used in WB test immediately to minimize protein degradation. -
All lanes : Anti-eNOS antibody [EPR26335-79] (ab300071) at 1/1000 dilution
Lane 1 : bEnd.3 (mouse brain endothelioma) whole cell fresh lysate
Lane 2 : RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell fresh 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: 133 kDa
Observed band size: 140 kDa why is the actual band size different from the predicted?This data was developed using ab300071, the same antibody clone in a different buffer formulation.
5% NFDM/TBST was used as blocking and diluting buffer.
Exposure time:
Lane 1: 10 seconds
Lane 2: 3 minutesNegative control:RAW264.7 (PMID: 18607531)
Lysates were freshly made and used in WB test immediately to minimize protein degradation. -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eNOS antibody [EPR26335-79] (BSA and Azide free) (AB300072)
This data was developed using ab300071, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded mouse spleen tissue labeling eNOS with ab300071 at 1/50 dilution (10.96 µg/ml), followed by a ready to use Leica DS9800 (BOND™ Polymer Refine Detection) kit. Positive staining on vascular endothelial cells of mouse spleen was observed. The section was incubated with ab300071 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 Leica DS9800 (BOND™ Polymer Refine Detection) kit.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins was used.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eNOS antibody [EPR26335-79] (BSA and Azide free) (AB300072)
This data was developed using ab300071, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded mouse colon tissue labeling eNOS with ab300071 at 1/50 dilution (10.96 µg/ml), followed by a ready to use Leica DS9800 (BOND™ Polymer Refine Detection) kit. Positive staining on vascular endothelial cells of mouse colon was observed (PMID: 12943528, PMID: 11479503). The section was incubated with ab300071 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 Leica DS9800 (BOND™ Polymer Refine Detection) kit.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins was used.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eNOS antibody [EPR26335-79] (BSA and Azide free) (AB300072)
This data was developed using ab300071, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded rat spleen tissue labeling eNOS with ab300071 at 1/50 dilution (10.96 µg/ml), followed by a ready to use Leica DS9800 (BOND™ Polymer Refine Detection) kit. Positive staining on vascular endothelial cells of rat spleen was observed. The section was incubated with ab300071 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 Leica DS9800 (BOND™ Polymer Refine Detection) was used.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins was used.
-
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eNOS antibody [EPR26335-79] (BSA and Azide free) (AB300072)
This data was developed using ab300071, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded rat stomach tissue labeling eNOS with ab300071 at 1/50 dilution (10.96 µg/ml), followed by a ready to use Leica DS9800 (BOND™ Polymer Refine Detection) kit. Positive staining on vascular endothelial cells of rat stomach was observed. The section was incubated with ab300071 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 Leica DS9800 (BOND™ Polymer Refine Detection) kit.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins was used.
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Immunocytochemistry/ Immunofluorescence - Anti-eNOS antibody [EPR26335-79] (BSA and Azide free) (AB300072)
This data was developed using ab300071, the same antibody clone in a different buffer formulation.
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized bEnd.3 (mouse brain endothelioma endothelialpolyoma middle T antigen transformed) cells lebeling eNOS with ab300071 at 1/50 dilution (10.96 µg/ml), followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2 µg/mL) (Green). Confocal image showing cytoplasm staining in bEnd.3 cell line. Negative control: RAW 264.7 (PMID: 18607531).
ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 (2.5µg/ml) dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 µg/mL) dilution.
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This data was developed using ab300071, the same antibody clone in a different buffer formulation.
Flow cyt (Intra) analysis of 4% paraformaldehyde fixed 90% methanol permeabilized RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage, Left) / bEnd.3 (Mouse brain endothelioma, Right) cells labeling eNOS with ab300071 at 1/500 dilution (0.1 µg) (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, ab150081) at 1/2000 dilution was used as the secondary antibody. Negative control: RAW 264.7 (PMID: 18607531).
Protocols
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.
Datasheets and documents
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Datasheet download
Certificate of Compliance
References (0)
ab300072 has not yet been referenced specifically in any publications.