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Fluorescent lifetime imaging (FLIM) is a powerful tool for visualizing physiological parameters in vivo. We present here a 3-dye strategy for mapping bioelectric patterns in living Xenopus laevis embryos leveraging the quantitative power of fluorescent lifetime imaging. We discuss a general strategy for disentangling physiological artifacts from true bioelectric signals, a method for dye delivery via transcardial injection, and how to visualize and interpret the fluorescent lifetime of the dyes in vivo.
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