Preparation of samples for Western blotting Sample Clauses

Preparation of samples for Western blotting. All samples were diluted to 3 µg of protein/µL using RIPA buffer before one volume of Laemmli sample buffer (Sigma-Xxxxxxx, Missouri, USA) was added (final protein concentration 1.5 µg/µL), mixed and heated to 100oC for 5 minutes using an Eppendorf heater (Grant Instruments, Shepreth, UK). This process denatures proteins and dissociates quaternary subunits by disrupting disulphide bridges and intramolecular bonds such as electrostatic interactions. As a consequence, protein separation during electrophoresis occurs primarily as function of protein size and not by quaternary structure.
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