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. 2006 Feb;50(2):587–595. doi: 10.1128/AAC.50.2.587-595.2006

FIG.3.

FIG.3.

(A) Differential uptake of TR-gentamicin and lucifer yellow in cells with the indicated genes mutated. A total of 500 μg/ml TR-gentamicin and 1 mg/ml lucifer yellow was added to cultures in the logarithmic-growth phase. Incubation at 30°C continued for 12 to 15 h. Yeasts were imaged using confocal microscopy with identical microscopic settings to enable direct comparison between strains. TR-gentamicin is shown in the top panel in red, DIC is shown in the middle panel, and lucifer yellow is shown in the bottom panel in yellow for each strain. Arrows identify the colocalization of TR-gentamicin and lucifer yellow in the vacuole. (B) Short-term uptake of TR-gentamicin and lucifer yellow. Overnight cultures were incubated with 500 μg/ml TR-gentamicin and 1 mg/ml lucifer yellow for 2 h. Imaging was conducted as for panel A. DIC image is shown in the left panel, with the TR-gentamicin shown in red in the middle panel. The overlay of all three channels was performed in Zeiss LSM Image Browser and imported into Adobe Photoshop, where color enhancement was performed. This process enabled the dimmer lucifer yellow to be visible when overlaid with the TR-gentamicin. Note the concentration of TR-gentamicin with lucifer yellow in the vacuole but also at distinct punctate locations in the cytoplasm.