The membranes were blocked with 5% skim milk for 1 hr at room temperature and incubated overnight at 4°C with primary antibodies against the β3 adrenergic receptors (1:1000, ab59685, Abcam), followed by incubation with appropriate horseradish peroxidase-linked secondary antibodies (1:4000) for 2 hr at room temperature. The bands on the blots were visualized using an enhanced chemiluminescence system (Amersham Bioscience, Buchinghamshire, UK), and densitometric analysis of the Western blots was conducted using VisionWorks LS, version 6.7.1.
Ab59685
Manufactured by Abcam
Sourced in United States
Ab59685 is a rabbit polyclonal antibody that targets an unspecified antigen. This antibody is suitable for use in various immunoassay applications, such as Western blotting and immunohistochemistry. The product is available in lyophilized format.
Automatically generated - may contain errors
Lab products found in correlation
Enhanced chemiluminescence system, by Cytiva (1 mentions)
L name, by Merck Group (1 mentions)
Ab4074, by Abcam (1 mentions)
Fv1000 confocal fluorescence microscope, by Olympus (1 mentions)
Ab6640, by Abcam (1 mentions)
Ab1426, by Abcam (1 mentions)
Sc 18096, by Santa Cruz Biotechnology (1 mentions)
Oapc00117, by Aviva Systems Biology (1 mentions)
Odyssey fc equipment, by LI COR (1 mentions)
Trans blot turbo system, by Bio-Rad (1 mentions)
Ab7120, by Abcam (1 mentions)
Ab9485, by Abcam (1 mentions)
4 protocols using ab59685
1
Quantifying Beta3 Adrenergic Receptor Expression
2
Sitagliptin-Induced Endothelial Function
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Sitagliptin (JANUVIA, 100 mg, Merck Sharp & Dohme, USA); streptozotocin, phenylephrine, isoprenaline, CL 316,243, L-NAME (Sigma, USA); anti-
ß 3 antibody (ab59685, Abcam, USA), anti-eNOS antibody (9572S, Cell Signaling Technology, USA), anti-phosphorylated eNOS (ser1177) antibody (9571S, Cell Signaling Technology, USA), anti-α-tubulin antibody (ab4074, Abcam, USA), antichicken antibody (29710, AnaSpec, USA), antirabbit antibody (7074, Cell Signaling Technology, USA).
3
Immunofluorescence Imaging of Adipocyte Markers
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Cells were grown on glass coverslips and fixed with 4% paraformaldehyde in PBS (pH 7.4) for 20 min at 4 °C. Cells were permeabilized with 0.3% Triton-X 100 in PBS for 30 min and blocked with PBS containing 3% goat serum and 1% bovine serum albumin for 1 h at room temperature. After incubation with rabbit anti-UCP-1 (EMD Millipore, #AB1426), Adrb3 (Abcam, #ab59685), and rat anti-F4/80 (Abcam, #ab6640) at 1:100 dilution overnight at 4 °C, the cells were washed with PBS and treated with goat anti-rabbit or anti-rat Alexa Fluor 488-conjugated and Alexa Fluor 594-conjugated secondary antibodies at 1:1000 dilution for 1 h. Cells were washed with PBS and incubated in 0.1 μg/ml Hochest (DAPI) for 30 min. After washing with PBS, coverslips were mounted onto microscope slides. Images were recorded using an Olympus FV1000 confocal fluorescence microscope.
4
Western Blot Analysis of Bladder Proteins
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Bladders were frozen in liquid nitrogen, pulverized in a mortar and dissolved using lysis buffer (62.5 mM Tris-HCL PH6.8, 2% SDS and 10% glycerol, with protease and phosphatase inhibitor cocktails added) (Shakirova et al., 2006) . 25 µg of protein was loaded in the wells of TGX Criterion gels (Any KD, BioRad). Protein was transferred to nitrocellulose membranes using the Trans-Blot Turbo system (BioRad). Membranes were incubated with primary antibodies against β3 adrenergic receptors (ab59685, Abcam, 1:2000), Avpr1a (sc-18096, Santa Cruz, 1:100), Glra1 (OAPC00117, Aviva systems biology, 1:1000), Nmbr (HPA026665, Sigma life science, 1:100) and Gapdh (ab9485, Abcam, 1:1000). Bands were visualized using horseradish peroxidaseconjugated secondary antibodies (SAB3700180, Sigma Aldrich, 1:5000, #7074, Cell Signaling, 1:5000, AB7120, Abcam, 1:5000) and West Femto chemiluminescence reagent (Pierce, Rockford, IL). Images were acquired using the LI-COR Odyssey Fc equipment (LI-COR Biosciences).
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