Isolation of the plant cytosolic fraction for proteomic analysis

The cytosol is the fluid portion of the cell that is not partitioned by membranes. It contains a highly diverse collection of substances and is central to many essential cellular processes ranging from signal transduction, metabolite production and transport, protein biosynthesis and degradation to stress response and defense. Despite its importance, only a few proteomic studies have been performed on the plant cytosol. This is largely due to difficulties in isolating relatively pure samples from plant material free of disrupted organelle material. In this chapter we outline methods for isolating the cytosolic fraction from Arabidopsis cell cultures and seedlings and provide guidance on assessing purity for analysis by mass spectrometry.