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. 2010 Feb 17;84(9):4124–4135. doi: 10.1128/JVI.02319-09

FIG. 5.

FIG. 5.

FRET by acceptor photobleaching demonstrates that ICP27 undergoes a head-to-tail intramolecular interaction but not an intermolecular interaction. (A and B) RSF cells were transfected with N-CFP/ICP27/C-YFP DNA. Cells were infected 24 h after transfection with 27-LacZ at an MOI of 10. Cells were fixed at 8 h after infection. (C) RSF cells were transfected with N-CFP/ICP27 and ICP27/C-YFP DNA. Cells were infected 24 h after transfection with 27-LacZ at an MOI of 10. Cells were fixed at 8 h after infection. CFP-YFP FRET by acceptor photobleaching was performed using an LSM 510 Meta confocal microscope as described by Karpova et al. (18) and detailed in Materials and Methods. The white circles indicate the specific area within each cell that was bleached at 514 nm. FRET efficiency, which is plotted to the right of the fluorescent images, was calculated as follows: FRET (%) = 1 − (donor before bleach − background before bleach)/(donor after bleach − background after bleach) × 100.