Figure 6.

A. Representative confocal slices. ES2 cells expressing ROR2-Emerald (green) on fibronectin coated glass +/− uniform Wnt5a signal. DAPI costain (blue).

B. Quantification of ratio of ROR2-Emerald (images in A) located on periphery of cell as as compared to total from confocal images. Integrated fluorescence intensity (relative fluorescence units (RFU)),n=90 per condition, student’s unpaired t-test, ****p<0.00001

C. ES2 cells expressing ROR2-Emerald loaded into middle chamber of gradient generating device and allowed to adhere to surface. Single cell migration tracked over 20 hours at 20 minute intervals, 40x magnification. Cell area quantified using brightfield imaging (white outline) and then bisected perpendicular to the movement vector. Representative images shown. White arrows represent areas of focal ROR2 intensity.

D,E. ROR2-Emerald (Green) intensity within leading quadrant of the cell was quantified and divided by the total green intensity within the cell to identify changes in ROR2 localization. Schematic of quantification shown in D, data quantified in E. ROR2 distribution within cell was quantified for cells that experience no gradient and those experiencing Wnt5a gradient (max concentration 5μg/mL). n>6 cells per device, 3 devices, unpaired t-test per time point (*, p<0.05; **, p<0.005).