Video microscopy was performed over a time period of 24 hours when using FCS as chemoattractant or twelve hours when using FaDu cells as attractant source. The time-lapse interval was ten minutes. Cell tracking was performed using the ImageJ software (National Institutes of Health, Bethesda, USA) plugin "Manual Tracking" (Fabrice Cordelières, Institut Curie, Orsay, France). Each experiment was repeated three times, completely independent from each other. On average 40-53 cells were tracked per experiment. In order to further analyse and evaluate chemotactical processes, we developed the ImageJ plugin, which is available for free as a download from the NIH ImageJ homepage. The "Chemotaxis and Migration Tool" provides different types of graphs and statistical tests based on the experimental data. Exported ASCII (mandatory format) tables from "manual tracking" were directly imported into the software tool, and the cell trajectories were all extrapolated to (x, y) = 0 at time 0 h.
For quantification of chemotaxis and migration, several values were generated to evaluate directed cell migration: the centre of mass, the forward migration indices in directions parallel and perpendicular to the direction of the gradient, and the Rayleigh [23 ] test as well as the cell velocity and directness, where the latter values provide indication for how straight cells move.
For quantification of chemotaxis and migration, several values were generated to evaluate directed cell migration: the centre of mass, the forward migration indices in directions parallel and perpendicular to the direction of the gradient, and the Rayleigh [23 ] test as well as the cell velocity and directness, where the latter values provide indication for how straight cells move.
Free full text: Click here
