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Live imaging of cellular internalization of single colloidal particle by combined label-free and fluorescence total internal reflection microscopy

Byrne, Gerard D.; Vllasaliu, Driton; Falcone, Franco H.; Somekh, Michael G.; Stolnik, Snjezana

Authors

Gerard D. Byrne

Driton Vllasaliu

Franco H. Falcone



Abstract

In this work we utilize the combination of label-free total internal reflection microscopy and total internal reflectance fluorescence (TIRM/TIRF) microscopy to achieve a simultaneous, live imaging of single, label-free colloidal particle endocytosis by individual cells. The TIRM arm of the microscope enables label free imaging of the colloid and cell membrane features, while the TIRF arm images the dynamics of fluorescent-labeled clathrin (protein involved in endocytosis via clathrin pathway), expressed in transfected 3T3 fibroblasts cells. Using a model polymeric colloid and cells with a fluorescently tagged clathrin endocytosis pathway, we demonstrate that wide field TIRM/TIRF coimaging enables live visualization of the process of colloidal particle interaction with the labeled cell structure, which is valuable for discerning the membrane events and route of colloid internalization by the cell. We further show that 500 nm in diameter model polystyrene colloid associates with clathrin, prior to and during its cellular internalization. This association is not apparent with larger, 1 μm in diameter colloids, indicating an upper particle size limit for clathrin-mediated endocytosis.

Journal Article Type Article
Publication Date Sep 24, 2015
Print ISSN 1543-8384
Publisher American Chemical Society
Peer Reviewed Peer Reviewed
Volume 12
Issue 11
Pages 3862-3870
APA6 Citation Byrne, G. D., Vllasaliu, D., Falcone, F. H., Somekh, M. G., & Stolnik, S. (2015). Live imaging of cellular internalization of single colloidal particle by combined label-free and fluorescence total internal reflection microscopy. Molecular Pharmaceutics, 12(11), 3862-3870. doi:10.1021/acs.molpharmaceut.5b00215
DOI https://doi.org/10.1021/acs.molpharmaceut.5b00215
Publisher URL https://pubs.acs.org/doi/10.1021/acs.molpharmaceut.5b00215

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