Anti-eNOS antibody [EPR26335-79]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eNOS antibody [EPR26335-79] (AB300071)

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.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-eNOS antibody [EPR26335-79] (AB300071)

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).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-eNOS antibody [EPR26335-79] (AB300071)

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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eNOS antibody [EPR26335-79] (AB300071)

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.

• WB

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Western blot - Anti-eNOS antibody [EPR26335-79] (AB300071)

5% NFDM/TBST was used as blocking and diluting buffer.

Exposure time :

Lane 1 : 10 seconds
Lane 2 : 3 minutes

Negative control : RAW264.7 (PMID : 18607531)
Lysates were freshly made and used in WB test immediately to minimize protein degradation.

All lanes:

Western blot - Anti-eNOS antibody [EPR26335-79] (ab300071) at 1/1000 dilution

Lane 1:

bEnd.3 (mouse brain endothelioma) whole cell fresh lysate at 20 µg

Lane 2:

RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell fresh lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 132 kDa,133 kDa

Observed band size: 140 kDa

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