Membrane localization, functional expression and specificity of bitter-responsive receptors in HEK293/15 cells. a-c, Confocal fluorescence images of HEK293/15 cells transfected with TAS2R16 (a), TAS2R10 (b) or rat T2R9 (c) DNA. The cell surface is visualized by plasma membrane glycoproteins detected with biotin-conjugated concanavalin A and avidin-conjugated Texas Red (red, left panels) and by receptors detected with a primary antibody against the HSV epitope and a Alexa488-conjugated secondary antibody (green, middle panels). Taste receptors that are located at the cell surface appear in yellow in the overlay (right panels). Optical sections were taken through the center of cells. Scale bars, 20 Ám. d-f, Mean ratio trace of the calcium response in single cells transfected with TAS2R16 (d), TAS2R10 (e) or rat T2R9 DNA (f), with 5 of 53 cells, 9 of 81 cells and 8 of 36 cells responding in d, e and f, respectively. Horizontal bars indicate the duration of application of 3 mM salicin (s), 200 ÁM strychnine (stry) and 10 ÁM cycloheximide (cyc). Similar results were obtained in two independent transfection experiments.

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