ImageJ intensity plots quantitatively confirm membrane and cortex positions of a putative bleb.

A single time point of a wild-type cell crawling under 0.5 percent agarose gel from Fig 2 with respective intensity plots of the signals along the drawn line (white line) for each channel is shown. Left, the LifeAct-GFP signal (dashed line) indicated the cortex’s position (white arrowhead) at 1.1 microns. Center, the RITC-Dextran signal (solid line) indicated the membrane’s position (black arrow) at 2.9 microns where the cell’s membrane ends and the agarose begins. Right, the merge of the two channels shows membrane detachment from the cortex by a distance of 1.8 microns.