Live Imaging of Cellular Internalization of Single Colloidal Particle by Combined Label-Free and Fluorescence Total Internal Reflection Microscopy

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

Research output: Journal article publicationJournal articleAcademic researchpeer-review

7 Citations (Scopus)

Abstract

� 2015 American Chemical Society. 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.
Original languageEnglish
Pages (from-to)3862-3870
Number of pages9
JournalMolecular Pharmaceutics
Volume12
Issue number11
DOIs
Publication statusPublished - 24 Sep 2015

Keywords

  • clathrin
  • endocytosis
  • evanescent wave microscopy
  • imaging systems
  • live cell imaging
  • total internal reflection fluorescence

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmaceutical Science
  • Drug Discovery

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