Improved method for measuring intracellular Ca++ with fluo-3 |
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| Authors: | G T Rijkers L B Justement A W Griffioen J C Cambier |
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| Institution: | Department of Immunology, University Hospital for Children and Youth, Het Wilhelmina Kinderziekenhuis, Utrecht, The Netherlands. |
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| Abstract: | The accuracy of flow cytometric measurement of intracellular calcium with fluo-3 is compromised by variation in basal fluorescence intensity due to heterogeneity in dye uptake or compartmentalization. We have loaded cells simultaneously with fluo-3 and SNARF-1. When SNARF-1 fluorescence is collected at approximately 600 nm, its intensity does not change upon cell activation. Furthermore, fluo-3 and SNARF-1 fluorescence signals exhibit a linear relationship. The ratio of fluo-3 to SNARF-1 eliminates a significant proportion of variation in fluorescence intensity caused by variation in fluo-3 uptake and thus can be used as a sensitive parameter for measuring changes in Ca2+]i. |
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