For Western blotting analyses, 50 μg of protein was resolved for each plasma sample by SDS-PAGE. Gels were transferred to 0.2 μm pore size Protran (BA83) nitrocellulose membranes (Whatman) in 25 mM Tris, 200 mM glycine. Membranes were blocked with 10% nonfat dry milk in PBS containing 0.1% Tween (PBST) for 1 hour at room temperature. Membranes were then probed with anti-FNDC5 (Irisin) rabbit polyclonal antibody (1:1000) (IN102, Adipogen) overnight at 4 °C, washed with PB ST and incubated with horseradish peroxidase-conjugated secondary antibodies (1:3000) (Sigma) followed by chemiluminescent (Perkin Elmer Life Sciences) detection. Recombinant mammalian irisin was obtained from Ember Therapeutics.
Protran ba83 nitrocellulose membranes
Manufactured by Cytiva
Protran (BA83) nitrocellulose membranes are a type of lab equipment used for various applications in scientific research. They are made of nitrocellulose and are designed to facilitate the immobilization and transfer of biomolecules such as proteins, nucleic acids, and other macromolecules. The core function of these membranes is to provide a suitable surface for these biomolecules to adhere to, enabling further analysis and processing.
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Lab products found in correlation
2 protocols using protran ba83 nitrocellulose membranes
1
Western Blotting Analysis of Irisin
2
Denaturing Cell Lysis and Immunoblotting
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Cells and spheroids were lysed in cold lysis buffer containing 3.7 M urea, 134.6 mM Tris, 5.4% SDS, 2.3% NP-40, 4.45% beta-mercaptoethanol, 4% glycerol, 6 mg/100 ml bromophenol blue (all from Sigma) according to Ehler et al. (1999 (link)). Cell lysates were further treated by repeated expelling through a fine needle and boiled for 5 min, then separated by SDS-PAGE using precast 4–20% gradient gels (BioRad) and blotted to Protran BA 83 nitrocellulose membranes (Whatman). Immunodetection was carried out after blocking in 5% milk in TBST (20 mM Tris base, pH 7.5, 150 mM NaCl, 0.05% Tween-20). For the visualization of signals, a LiCor Odyssey infrared imaging system (LiCor) was used with secondary antibodies coupled to Alexa700 and Alexa790 fluorescent dyes (ThermoFisher).
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