A range of techniques have been used to measure the concentration of cytosolic-free Ca2+ ([Ca2+]cyt) in plant cells. Fluorescent Ca2+-sensitive indicators have been used extensively to measure plant [Ca2+]cyt and a number of techniques are available for loading these into plant cells. Here we describe a method for measuring [Ca2+)cyt in the guard cells of the model plant species Commelina communis by ratio photometry and imaging techniques using the ratiometric fluorescent Ca2+-sensitive indicator fura-2