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autoFISH - a modular toolbox for sequential smFISH experiments

Weber, C.; Defard, T.; Lelek, M.; Laporte, H.; Mallick, A.; Londono-Vallejo, J.-A.; Walter, T.; Fouillade, C.; Gariboldi, M. I.; Muller, F.

2024-11-15 molecular biology
10.1101/2024.11.14.623566 bioRxiv
Show abstract

Fluorescence in situ hybridization (FISH) allows for spatial and quantitative profiling of gene expression by visualizing individual RNA molecules. Here, we introduce automated FISH (autoFISH), a comprehensive toolbox to conduct automated single molecule FISH (smFISH) experiments that is both cost-effective and versatile. This includes detailed plans for constructing the necessary equipment, open-source software for control, reliable experimental protocols, and analysis workflows based on our FISH-quant analysis package. Validation experiments with both cell lines and tissue samples confirmed the systems robustness. We demonstrate standard and amplified smFISH, along with a modified protocol for tissue clearing that enhances nuclear retention while preserving background reduction efficiency.

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