Supporting Information

Files in this Data Supplement:

Supporting Figure 7
Supporting Materials and Methods
Supporting Figure 8
Supporting Table 1
Supporting Table 2
Supporting Figure 9




Supporting Figure 7

Fig. 7. Automated wound-healing assay device. (1) Sterilized pipette tips are inserted into an aluminum block and suspended on a vertical tracking arm. The tip block is raised against a top plate to stabilize the tip position and prevent any movement upon scratching. The 384-well clear bottom tissue culture plates are placed on a level platform below the aluminum block. Once the equipment is initiated, the aluminum block is automatically lowered to a point at which the pipette tips touch the bottom of each of the 384 wells. (2) With the pipette tip block engaged, the platform is shifted » 3 mm (well diameter = 3.70 mm) by hydraulic pressure, resulting in uniform "scratches" in each of the 384 wells. After scratching, the tip block is raised up from the plate, and the plate holder returns to the start position to allow manual plate switching by the user.

Supporting Figure 8

Fig. 8. Reproducibility of the 384-well scratch assay. The quantitative migration score of each well of a 384-well plate was first averaged across three replicate plates. Scores from each individual run were then compared to the average (represented by the different colors in "single score"), resulting in r² values >0.87 for all comparison.

Supporting Figure 9

Fig. 9. Illustration of the qualitative migration scoring algorithm. (a) Raw data consist of one grayscale image per well. Bright regions represent cells, and black regions represent background. (b) The grayscale image is converted to a binary black and white mask image. To avoid taking any potential unscratched regions into account, the left and right 25 percent of the original image are cropped out. (c) The program calculates the number of white pixels for every row in the image, and the resulting curve represents cell density as a function of vertical location. The green line represents the average number of white pixels outside the scratch window. The purple dashed area A_s is proportional to the number of cells being removed by the scratch. The yellow area A_m is proportional to the number of cells migrating back into the scratch zone. A score close to 1 is assigned to cells with high motility, and a score close to 0 is assigned to cells with low motility. (d) The scratch score S is iteratively calculated for every possible scratch center within a given range. The minimal possible S score is reported.