To begin, pour 100 milliliters of the extraction buffer into a one liter plastic beaker. Determine the weight of the buffer and vessel, W1.Take the porcine brain out of the transport vessel. Using fingers and moderate force, remove the cerebellum, fat, big chunks of white matter, meninges, and blood vessels from the brain.
Place the stripped brain into the beaker containing 100 milliliters of cold extraction buffer, and determine its weight, W2.Add the calculated amount of extraction buffer to the beaker. Transfer the brain tissue with extraction buffer into the pre-chilled kitchen blender and process in four to eight short three-second pulses. Then using a high-speed dispersion homogenizer, process the partially homogenized tissue for one minute.
After that, leave the suspension on ice for three to five minutes while occasionally mixing with a spoon. Pour the cell suspension into pre-chilled ultracentrifugation tubes and centrifuge the tubes at 50, 000g for one minute at four degrees Celsius. Retain the supernatant and discard the pellet.
