Thank you for your comment!

I measure the density of the sheet two different ways. When I track individual cells or measure the area of the cell sheet (Figs 5 and 6) I am staining the cells with a dye called CMFDA. This allows me to identify the centroid of the cell by its peak fluorescence intensity (I used a software called Imaris from Bitplane). In this case, as I have identified the positions of the cells, I just count the total number of positions in the first time frame, and divide by the area that they occupy. The second way, is when measuring retraction of the sheet after denudation or blebbistatin treatment (Fig 3, and Supplemental Figures), I count by hand. Since I do not stain the cells, but rather image under brightfield, I count the nuclei in the first time frame and again divide by the area that they occupy (there are almost never any multi-nucleate cells). However, this is impossible when the density is very high and the nuclei are hard to identify (which is why I do not even assign a number to the highest density, but state the density is >3000 cells/mm2). Also, there is more error in counting by hand, so I choose to broadly classify density into low, intermediate, and high.

I hope this helps!