Fluo-4 AM

A fluorescent calcium indicator

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Background

Fluo-4 acetoxymethyl ester (Fluo-4 AM) is a cell-permeable fluorescent calcium indicator.¹ It is cleaved by intracellular esterases to release Fluo-4, which binds to calcium with a K_d value of 345 nM and displays excitation/emission maxima of 490/520 nm, respectively. Fluo-4 AM has been used to measure intracellular calcium levels in live cells by fluorescent microscopy or flow cytometry.^1,2

1.Gee, K.R., Brown, K.A., Chen, W.N., et al.Chemical and physiological characterization of fluo-4 Ca2+ - indicator dyesCell Calcium27(2)97-106(2000) 2.Vines, A., McBean, G.J., and Blanco-Fernández, A.A flow-cytometric method for continuous measurement of intracellular Ca2+ concentrationCytometry A.77(11)1091-1097(2010)

Protocol

Cell experiment:

For measuring fluorescence from cells in suspension, dilutions to 2 to 3×106 cells are made, from cultures of rat basophilic leukemia (RBL) cells. Cells are incubated in suspension in 1 mM dye (including Fluo-4 AM) for 30 min at 37°C. Cell suspensions are then transferred to cuvets for measurements of fluorescence emission intensity by spectrofluorometer[1].

参考文献:

[1]. Gee KR, et al. Chemical and physiological characterization of fluo-4 Ca(2+)-indicator dyes. Cell Calcium. 2000 Feb;27(2):97-106.