This contains the pipelines and data processing used for analyzing data from the Fluorecent actin stainging high-content (FAS-HCS) assay.
The assay was originally described by Stuart Knutton et al in 1989. The authors show that so called attaching and effacecing lesions caused by various pathogens can be visualized with flourecent phallotoxins. At that time it had been established that these pathogens lesion the brush boder microvillous membrane, and that they also do it in human tissue culture cell lines. Electron microscopy demonstrated dense concentrations of microfilaments present in the apical cytoplasm beneath the attached bacteria. Using fluorecein conjugated to phatllotoxins - toxins that strongly bind to actin filaments - the authors demonstrated that the lesions are rich in cytoskeletal actin and created a new way to visualize the phenotype.
A high throughput method to collect this data can be created by using fluorecent bacteria (Pylkkö 2021).
literature:
Knutton, S., T. Baldwin, P. H. Williams, and A. S. McNeish. 1989. “Actin Accumulation at Sites of Bacterial Adhesion to Tissue Culture Cells: Basis of a New Diagnostic Test for Enteropathogenic and Enterohemorrhagic Escherichia Coli.” Infection and Immunity 57 (4): 1290–98.
Pylkkö, Tuomas, Polina Ilina, and Päivi Tammela. 2021. “Development and Validation of a High-Content Screening Assay for Inhibitors of Enteropathogenic E. Coli Adhesion.” Journal of Microbiological Methods, March, 106201. https://doi.org/10.1016/j.mimet.2021.106201.