Comparative profiling of cellular gait on adhesive micropatterns defines statistical patterns of activity that...

Cell dynamics are powered by patterns of activity, but it is not straightforward to quantify these patterns or compare them across different environmental conditions or cell-types. Here we digitize the long-term shape fluctuations of metazoan cells grown on micropatterned fibronectin islands to define and extract statistical features of cell dynamics without the need for genetic modification or fluorescence imaging. These shape fluctuations generate single-cell morphological signals that can be decomposed into two major components: a continuous, slow-timescale meandering of morphology about an average steady-state shape; and short-lived "events" of rapid morphology change that sporadically occur throughout the timecourse. By developing statistical metrics for each of these components, we used thousands of hours of single-cell data to quantitatively define how each axis of cell dynamics was impacted by environmental conditions or cell-type. We found the size and spatial complexity of the micropattern island modulated the statistics of morphological events-lifetime, frequency, and orientation-but not its baseline shape fluctuations. Extending this approach to profile a panel of triple negative breast cancer cell-lines, we found that different cell-types could be distinguished from one another along specific and unique statistical axes of their behavior. Our results suggest that micropatterned substrates provide a generalizable method to build statistical profiles of cell dynamics to classify and compare emergent cell behaviors.