Improvement in Transient Agarose Spot (TAS) Cell Migration Assay: Microplate-Based Detection and Evaluation

Veres-Székely, Apor and Szász, Csenge and Pap, Domonkos and Bokrossy, Péter and Lenzinger, Dorina and Visnovitz, Tamás and Mihály, Judith and Pálmai, Marcell and Varga, Zoltán and Őrfi, László and Szabó, Attila and Vannay, Ádám and Szebeni, Beáta (2025) Improvement in Transient Agarose Spot (TAS) Cell Migration Assay: Microplate-Based Detection and Evaluation. INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES, 26 (12). No. 5584. ISSN 1661-6596

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Abstract

Collective cell migration is crucial in various biological processes, including tumor progression and metastasis. The widely used scratch assay (wound healing assay) has limitations in throughput, reproducibility, and data analysis. To overcome these challenges, we previously developed the Transient Agarose Spot (TAS) assay, which enhanced assay precision and reproducibility. In this study, we present an improved microplate-based TAS assay. By using a microplate reader, we automated data acquisition, enabling the detection of cell migration in a 96-well plate format with greater throughput and accuracy. The new method applies Hoechst staining to label viable cells, providing a stable signal for kinetic analysis without compromising cell viability. We validated this approach with fluorophore-expressing cancer cells and demonstrated its ability to monitor dose-dependent effects of fetal bovine serum on cell migration. Additionally, we applied the microplate-based TAS assay to assess the anti-migratory effects of kinase inhibitors and mesenchymal stem cell-derived extracellular vesicles (EVs) on lung cancer cells. The assay accurately quantified migration inhibition and revealed the concentration-dependent effects of EVs, highlighting their potential as therapeutic agents. This microplate-based TAS assay provides a scalable, efficient, and cost-effective platform for high-throughput screening of cell migration and drug discovery, offering a robust alternative to traditional microscopy-based methods.

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