This protocol describes a method to assess concentrations of free cytoplasmic calcium, [Ca2+], for cultured adherent RAW 264.7 cells in an 8-well coverglass. This objective is accomplished using the Ca2+-sensitive fluorescent dye, fura-2
acetoxymethyl (AM), which permeates cell membranes as an ester and is hydrolyzed in the cell to its Ca2+-sensitive acidic form.