Abstract
The eukaryotic cytoskeleton plays essential roles in cell signaling and trafficking, broadly associated with immunity and diseases in humans and plants. To date, most studies describing cytoskeleton dynamics and function rely on qualitative/quantitative analyses of cytoskeletal images. While state-of-the-art, these approaches face general challenges: the diversity among filaments causes considerable inaccuracy, and the widely adopted image projection leads to bias and information loss. To solve these issues, we developed the Implicit Laplacian of Enhanced Edge (ILEE), an unguided, high-performance approach for 2D/3D-based quantification of cytoskeletal status and organization. Using ILEE, we constructed a Python library to enable automated cytoskeletal image analysis, providing biologically interpretable indices measuring the density, bundling, segmentation, branching, and directionality of the cytoskeleton. Our data demonstrated that ILEE resolves the defects of traditional approaches, enables the detection of novel cytoskeletal features, and yields data with superior accuracy, stability, and robustness. The ILEE toolbox is available for public use through PyPI and Google Colab.