Ratiometric analysis of calcium mobilization
One of the earliest events in lymphocyte activation is an increase in the concentration of cytosolic free Ca2+. Antigen recognition, chemotaxis, apoptosis and cell cycle progression are all associated with changes in cytosolic free Ca2+ levels. The kinetics, subcellular localization, and degree of Ca2+ mobilization can be assessed with fluorescent Ca2+ indicators. Ratiometric analysis of Ca2+ mobilization reduces errors due to variations in cell size and shape, indicator loading and instrument variability that can otherwise limit the usefulness of fluorescent Ca2+ indicators. The purposes of this paper are to summarize key characteristics of some useful Ca2+ -sensitive fluorescent dyes (Indo-1, Fura-2, Fluo-3, Fluo-4 and Fura Red) and to review published methods for the optimization of Ca2+ mobilization analysis with ratiometric indicators. Analysis of Ca2+ mobilization provides a rapid, quantitative readout of lymphocyte activation. © 2000 Elsevier Science Inc. All rights reserved.