Fig. 4.

CTP forms a practical workflow for managing cell segmentation and tracking and can feed outputs into a variety of tools for statistical analysis. CTP runs in Icy under ‘plugins’. Users define segmentation parameters for processing of datasets by HK-Means. Outputs, including images and measurements of cells were saved by ‘Time-Saver’ into two files. Objects are tracked by the Keyhole Tracker program in MATLAB Runtime then imported into the Icy plugin Track Manager for editing. The Milkshake plugin then collates information from tracking with values of cell shape (from file ROI+Labels) to produce a tabulated sheet of data. This can be analysed using a MATLAB script, CTP2R, which can normalise data to time-point 0 (t₀), generate values of directional autocorrelation, mean squared displacement (MSD) and select a region of interest (ROI), and average parameter values over time. Output files in CTP are shown in italics, modified Icy plugins are shown in a grey box, new scripts and plugins are shown in boxes, and plugins shown in bold indicate user input is required.