Abstract
This chapter examines the properties of Clomeleon, a genetically encoded fluorescent indicator that permits non-invasive, spatiotemporally resolved optical recordings of steady-state as well as dynamic changes of [Clˉ]i. It also reviews the application of Clomeleon to [Clˉ]i measurements in neurons, the cell type that interests us the most. Clomeleon is a fusion protein consisting of the cyan fluorescent protein (CFP) and the topaz variant of yellow fluorescent protein (YFP). A linker of 24 amino acid residues containing an rTEV (recombinant tobacco etch virus protease) cleavage site connects the C-terminus of CFP to the N-terminus of YFP. This linker keeps the two fluorescent proteins in close proximity, permitting fluorescence resonance energy transfer (FRET) between CFP and YFP. Thus, illumination of Clomeleon with blue light excites CFP, resulting in radiationless FRET to YFP.
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