Once samples have been bound to the metal affinity resin, the procedures for purification and desalting are clearly outlined in the respective manufacturers’ manuals. To prepare the cell pellets for binding to the resin, the cells will be passed through the Microfluidizer in the Mehl Core Lab followed by centrifugation of the lysed cells. Alternatively, cells must be sonicated and centrifuged according to the general procedure provided later in this manual. Note that throughout the purification process, the samples should be kept on ice to prevent less stable proteins from unfolding.
Of the three types of cell pellets being stored at -80 °C, purify at least one pellet of each type. Save the remaining pellets at -80°C. More protein expressions can be done later as needed. Note, however, that at least three days’ notice is required to prepare more non-inducing starters.