Western Blot Quantification Protocol

Proteins separated by electrophoresis were transferred onto PVDF (polyvinylidene difluoride) membrane (Immobilon FL 0.45 µm, Merck) by semi-dry electroblotting (0.8 mA/cm², 1 h) using Transblot SD apparatus (Bio-Rad). Immunodetection was performed as described in [15 (link)]. Primary and secondary antibodies used for immunodetection are listed in Table S1. Signal detection was performed using fluorescence scanner Odyssey (LI-COR Biosciences, Lincoln, NE, USA). Signals detected by scanner were quantified by software AIDA Image Analyzer (Raytest Isotopenmessgeräte GmbH, Germany).

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Čunátová K., Reguera D.P., Vrbacký M., Fernández-Vizarra E., Ding S., Fearnley I.M., Zeviani M., Houštěk J., Mráček T, & Pecina P. (2021). Loss of COX4I1 Leads to Combined Respiratory Chain Deficiency and Impaired Mitochondrial Protein Synthesis. Cells, 10(2), 369.

Publication 2021

Immobilon FL 0.45 µm Transblot SD apparatus AIDA Image Analyzer

Antibodies Electrophoresis Fluorescence Immobilon Membrane Polyvinylidene difluoride Proteins Signal detection

Corresponding Organization : Czech Academy of Sciences, Institute of Physiology

Other organizations : Charles University, MRC Mitochondrial Biology Unit, University of Cambridge

Top 5 similar protocols

Variable analysis

independent variables

Proteins separated by electrophoresis

dependent variables

Signal detected by the fluorescence scanner Odyssey
Signals quantified by the AIDA Image Analyzer software

control variables

PVDF (polyvinylidene difluoride) membrane (Immobilon FL 0.45 µm, Merck)
Semi-dry electroblotting (0.8 mA/cm^2, 1 h) using Transblot SD apparatus (Bio-Rad)
Immunodetection performed as described in [15]
Primary and secondary antibodies used for immunodetection listed in Table S1

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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