Two-Dimensional Semi-Denaturing Agarose Gel Electrophoresis: A Technique to Separate Polymorphic Amyloids Fibers Based on Size Heterogeneity
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To prepare the gel, add 2 grams of agarose powder to 200 milliliters of TAE buffer in a glass beaker. Then, heat the beaker in a microwave to melt the agarose.
Next, add 1 milliliter of 20% SDS to a final concentration of 0.1%. Gently swirl the beaker.
Next, pour the liquid agarose on a 15 cm x 14 cm gel slab. To remove any air bubbles, use a 1-milliliter pipette. Then, place a 20-well comb on top of the gel.
Next, add approximately 60 micrograms of whole-cell lysate in the far-right lane of the gel. Run the gel at 60 volts for about four hours, using the TAE buffer containing 0.1% SDS, as the running buffer.
To run the gel in the second dimension, carefully rotate the gel by 90 degrees counter-clockwise. Then, run the jail at 60 volts for about four hours.
