Description

Time-lapse TIRF visualization of GFP-tagged EB1 showed bright foci of EB1-GFP moving away from microtubule seeds locations corresponding to the growing microtubule ends, confirming that EB1-GFP is capable of tip-tracking growing microtubule ends without the need of any protein binding partners. Tip-tracking was observed at both the plus and minus ends of growing microtubules, but no end localization was seen for microtubules that were not growing. Bright EB1-GFP foci were not observed at the microtubule ends during periods of microtubule depolymerization. The above findings confirm previous results that the tip-tracking of EB1 relies on the process of microtubule growth. Images were taken at 5 second intervals. Microtubules were (red, imaged by EPI fluorescence) with 24 µM unlabeled tubulin, 50 nM EB1-GFP (green, imaged by TIRF) in Imaging Buffer (see Methods in corresponding paper) with additional 60 mM KCl. Video-playback is 50× times real time. Area size is 45 µmx31 µm. Video corresponds to supplemental video 1 in PLoS One. 2009 Oct 23;4(10):e7585.