Isolation of Apoplast.

The apoplast can be described as the soluble fraction of the extracellular space of plant tissue, and it plays an important role in signaling, nutrient transport, and plant-pathogen interactions. In this protocol, we describe a method where leaves are infiltrated with phosphate buffer under vacuum. The apoplast can then be extracted by centrifugation and simultaneously collected in a protease inhibitor solution. Using this protocol, typically 3 μg of apoplastic proteins can be obtained in a volume of 300 μL from five potato leaflets, with minimal contamination by non-apoplastic proteins.