Lysing Yeast Cells for Immunoprecipitation Using a Coffee Grinder

Abstract

This protocol describes the freezing of yeast in liquid nitrogen (LN2) to form small "beans" that can be ground using a simple propeller-blade coffee grinder. The method is ideally suited for lysate preparations from larger yeast cultures ranging from 50 mL to 5 L and displays the advantage that samples remain cold during the preparative steps. Cells are cultured and collected by centrifugation while in log phase, and the resultant cell pellets are mixed with deionized distilled water and dropped into LN2 to form small frozen beans. Before the freezing process, it is imperative to keep all cell pellets at 4°C on ice. The frozen yeast beans are ground by using a simple kitchen coffee grinder, and the yeast powder is collected for immediate lysis or storage at -80°C for subsequent use. Protective clothing and safety glasses should be worn at all times when working with liquid nitrogen. Plasticware may shatter upon repeated cooling in liquid nitrogen, and appropriate care should be taken.