Tissue samples (20 μg/lane) were run in 7.5% Tris-Glycine gels and transferred onto nitrocellulose membranes (sc-3724, Santa Cruz Biotechnology). The nitrocellulose membranes were blocked in 10% (w/v) semi fat dry milk in TBS (Tris 0.01 M, NaCl 0.15 M, pH 7.4) for 1 h at room temperature and were incubated with primary antibodies overnight followed by rinses and an incubation with anti-rabbit (#7074, Cell Signalling Technology, Bioké) or anti-mouse (A6782, Sigma) immunoglobulin conjugated to horseradish peroxidase. After several rinses, the membranes were incubated with SuperSignal West Pico Substrate (Pierce) and were exposed to an X-ray film (Pierce) or to a DARQ-7 CCD cooled camera (Vilber-Lourmat) in a SOLO 4S WL system. Levels of optical density (OD) of protein signals were estimated by densitometry analysis using the NIH ImageJ program. Anti-β-actin immunoblots were used to normalize protein loading.
Darq 7 ccd cooled camera
Manufactured by Vilber
The DARQ-7 is a CCD-based camera designed for scientific and industrial applications. It features a cooled CCD sensor to minimize dark current and thermal noise. The camera provides high-resolution image capture capabilities, making it suitable for a variety of imaging tasks.
Automatically generated - may contain errors
2 protocols using darq 7 ccd cooled camera
1
Western Blot Protein Analysis
2
Western Blot Protein Detection
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Protein concentrations were estimated by the Bradford method (Bio-Rad). Samples (20 µg/lane) were run in 10 % or 15 % of Tris-glycine gels depending on the molecular weight of the protein of interest and transferred onto PVDF membranes (Bio-Rad). The PVDF membranes were blocked in 10 % (w/v) semi fat dry milk in TBS (Tris-buffered saline, 20mM Tris pH 7.6, 150 mM NaCl) for 1 h at room temperature and were incubated with primary antibodies overnight at 4°C. After three times of rinses with TBS-T (TBS supplemented with 0.05% Tween 20), the membranes were incubated with anti-rabbit (G-21234, Invitrogen) or anti-mouse (G-21040, Invitrogen) immunoglobulin conjugated with horseradish peroxidase. After three rinses with TBS-T, the membranes were incubated with SuperSignal West Pico Substrate (Pierce) and were exposed to an X-ray film (Pierce) or to a DARQ-7 CCD cooled camera (Vilber-Lourmat) in a SOLO 4S WL system. Levels of proteins were estimated by densitometry analysis using the NIH Image J program, and adjusted for protein loading based on western blots performed with an anti-actin antibody.
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