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. 2004 Jul 5;166(1):27–36. doi: 10.1083/jcb.200401056

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Copyright © 2004, The Rockefeller University Press

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Figure 3. — Dynamic measurements of the GFP-CSB nuclear mobility by combined FLIP/FRAP analysis. (A) Combined FLIP/FRAP analysis was performed by bleaching at one pole of the nucleus and simultaneously monitoring the fluorescent recovery at the bleached (FRAP) and opposite (FLIP) poles of the cell. After bleaching, the FRAP curve shows a drop in fluorescent intensity followed by a recovery of fluorescence and the FLIP curve shows a slow decrease of fluorescent intensity due to redistribution of the bleached molecules. The relative intensities of FLIP and FRAP were subtracted and plotted (y axis) against the relative redistribution time of untreated cells (x axis). (B) Combined FLIP/FRAP experiment of untreated cells (blue circles; n = 10) and H8-treated cells (red diamonds; n = 10). H8-treated cells display a decreased relative redistribution time as compared with untreated cells. (C) Combined FLIP/FRAP experiment of untreated cells (blue circles; n = 10) and Actinomycin D–treated cells (red diamonds; n = 10). Actinomycin D treatment results in an increased relative redistribution time as compared with untreated cells. (D) Combined FLIP/FRAP experiments at different temperatures (27°C, red triangles; 32°C, orange diamonds; and 37°C, blue circles). At low temperatures the relative redistribution time is increased. (E) Combined FLIP/FRAP experiment of untreated cells (blue circles, n = 10) and azide-treated cells (red diamonds; n = 10). Azide-treated cells display a decreased relative redistribution time as compared with untreated cells.