Ca2+ imaging in single living cells: theoretical and practical issues.
The measurement of intracellular calcium ion concentrations [( Ca2+]i) in single living cells using quantitative fluorescence microscopy draws from a diverse set of disciplines, including cellular biology, optical physics, statistics and computer science. Over the last few years, we have devised and built a number of systems for measuring [Ca2+]i with Fura-2, and have applied them in the exploration of a wide range of biological processes controlled by Ca2+. In this report we discuss these systems and their advantages and limitations. We also describe the theoretical and practical problems associated with using Fura-2 to measure [Ca2+]i, and the solutions that we, and others, have developed to overcome them. The approaches described should provide useful guidance for others interested in imaging [Ca2+] distribution in living cells. The factors that limit current methods are discussed, and areas for future development are highlighted.
Department of Physiology, University of Massachusetts Medical School, Worcester.
E D
ED
Moore
P L
PL
Becker
K E
KE
Fogarty
D A
DA
Williams
F S
FS
Fay
D000818:Animals; D001572:Benzofurans; D002118:Calcium / Q000032:analysis*; D002478:Cells, Cultured; D005456:Fluorescent Dyes; D016257:Fura-2; D006801:Humans; D008856:Microscopy, Fluorescence
D001572:Benzofurans; D005456:Fluorescent Dyes; D002118:Calcium; D016257:Fura-2