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This video tutorial demonstrates how to prepare a western blot membrane for chemiluminescent detection. After antibody incubation and washing, the membrane is ready for enhanced chemiluminescent (ECL) substrate. Use 7ml of substrate for a mini-sized membrane and 12ml for a medium-sized one for optimal results. A clean sheet protector is recommended to reduce substrate volume. Incubate for five minutes to ensure uniform coating and displacement of wash buffer. Membranes can be stored in the sheet protector for imaging.