Automated Multi-Peak Tracking Kymography (AMTraK): A Tool to Quantify Sub-Cellular Dynamics with Sub-Pixel Accuracy

Area of Science:

• Cell Biology
• Biophysics
• Microscopy

Background:

• Kymographs are essential for analyzing spatio-temporal dynamics in microscopy time-series data.
• Manual quantification of kymographs is labor-intensive and prone to subjectivity.
• Existing tools often lack automated quantification capabilities for complex dynamics.

Purpose of the Study:

• To develop and validate a novel software tool, Automated Multi-Peak Tracking Kymography (AMTraK), for automated quantification of kymographs.
• To enable objective and reproducible analysis of sub-cellular dynamics from fluorescence microscopy time-series.
• To track and quantify complex events like merging and splitting of dynamic features.

Main Methods:

• AMTraK utilizes peak information and distance minimization for automated tracking and quantification of kymographs.
• The software integrates a branch-point detection method to identify track merging and splitting events.
• It processes fluorescence time-series data, tracking contours based on intensity and gradient peaks.

Main Results:

• AMTraK demonstrated sub-pixel positional accuracy in tests with synthetic images.
• The software accurately quantified sub-cellular dynamics in E. coli cell division, microtubule transport, and vesicle dynamics.
• Analysis of previously manually quantified data reproduced reported parameters, validating AMTraK's performance.

Conclusions:

• AMTraK provides an objective, automated, and reproducible method for analyzing kymographs from fluorescence microscopy.
• The software facilitates the quantitative study of diverse in vitro and in vivo sub-cellular dynamics.
• AMTraK significantly advances the capabilities for analyzing spatio-temporal data in cell biology.