Empowering High-Throughput High-Content Analysis of Microphysiological Models: Open-Source Software for Automated...

Area of Science:

• * Cellular and Molecular Bioengineering
• * Biomedical Engineering
• * Computational Biology

Background:

• * Existing tools for analyzing dynamic cell behaviors in microphysiological models are often inaccessible or proprietary.
• * High-throughput analysis of endothelial tube formation and cell invasion is crucial for drug discovery.
• * Non-confocal microscopy is a widely available imaging technique that could benefit from advanced analysis tools.

Purpose of the Study:

• * To develop an open-source Python-based software for automated analysis of dynamic cell behaviors in microphysiological models using non-confocal microscopy.
• * To address the gap in accessible tools for high-throughput analysis of endothelial tube formation and cell invasion.
• * To facilitate the rapid assessment of drug sensitivity in vitro.

Main Methods:

• * Annotated over 1000 Z-stacks of cancer and endothelial cell co-culture models.
• * Trained machine learning models (ResNet-50, U-Net Xception-style) for cell coverage, invasion depth, and microvessel dynamics quantification.
• * Utilized focus stacking, Gaussian mixture models, DisPerSE algorithm, and graph analysis for comprehensive feature extraction.

Main Results:

• * Developed software accurately measures cell coverage, cancer invasion depth, and microvessel length.
• * Achieved drug sensitivity IC50 values with 95% confidence, comparable to manual calculations.
• * Significantly reduced image processing time from weeks to hours, demonstrating high-throughput capability.

Conclusions:

• * The free, open-source software provides an automated solution for quantifying 3D cell behavior in microphysiological models using non-confocal microscopy.
• * Offers a versatile alternative to standard confocal microscopy and proprietary software for the bioengineering community.
• * Software is available on GitHub, promoting accessibility and further development.