Movie 3

Epifluorescence time-lapse of a Bra-GFP embryoid body, where Bra onset occurred at the bottom. The EB is imaged between 24 and 90 hrs from aggregation, where at 24 hrs it was transferred to a microwell. Top row: left - brightfield imaging of the EB with its encompassing perimeter; center - Bra-GFP; right – overlay of brightfield and GFP-positive pixels (magenta). Bottom row: left - total Bra-GFP+ pixels vs. time point (time interval between points – 30 minutes). Blue line - raw data; red dashed line: alpha filter smoothing; yellow line: Bra onset threshold defined as 500 GFP+ pixels; center – EB radius vs. timepoint. Noise at higher time points is due to manual estimation of radius; right – snapshot of Bra-GFP onset frame.