title: Quantification of gold nanoparticle cell uptake under controlled biological conditions and adequate resolution.
abstract: OBJECTIVE
  We examined cellular uptake mechanisms of fluorescently labeled polymer-coated gold nanoparticles (NPs) under different biological conditions by two quantitative, microscopic approaches.
  
  
  METHODS
  Uptake mechanisms were evaluated using endocytotic inhibitors that were tested for specificity and cytotoxicity. Cellular uptake of gold NPs was analyzed either by laser scanning microscopy or transmission electron microscopy, and quantified by means of stereology using cells from the same experiment.
  
  
  RESULTS
  Optimal inhibitor conditions were only achieved with chlorpromazine (clathrin-mediated endocytosis) and methyl-β-cyclodextrin (caveolin-mediated endocytosis). A significant methyl-β-cyclodextrin-mediated inhibition (63-69%) and chlorpromazine-mediated increase (43-98%) of intracellular NPs was demonstrated with both imaging techniques, suggesting a predominant uptake via caveolin-medicated endocytois. Transmission electron microscopy imaging revealed more than 95% of NPs localized in intracellular vesicles and approximately 150-times more NP events/cell were detected than by laser scanning microscopy.
  
  
  CONCLUSIONS
  We emphasize the importance of studying NP-cell interactions under controlled experimental conditions and at adequate microscopic resolution in combination with stereology.
authors.affiliation: Adolphe Merkle Institute, University of Fribourg, Marly, Switzerland.
authors.forename: Barbara
authors.initials: B
authors.lastname: Rothen-Rutishauser
authors.forename: Dagmar A
authors.initials: DA
authors.lastname: Kuhn
authors.forename: Zulqurnain
authors.initials: Z
authors.lastname: Ali
authors.forename: Michael
authors.initials: M
authors.lastname: Gasser
authors.forename: Faheem
authors.initials: F
authors.lastname: Amin
authors.forename: Wolfgang J
authors.initials: WJ
authors.lastname: Parak
authors.forename: Dimitri
authors.initials: D
authors.lastname: Vanhecke
authors.forename: Alke
authors.initials: A
authors.lastname: Fink
authors.forename: Peter
authors.initials: P
authors.lastname: Gehr
authors.forename: Christina
authors.initials: C
authors.lastname: Brandenberger
mesh_terms: D002460:Cell Line; D006046:Gold / Q000032:analysis / Q000737:chemistry*; D006801:Humans; D008168:Lung / Q000737:chemistry* / Q000648:ultrastructure*; D018613:Microscopy, Confocal / Q000379:methods*; D008854:Microscopy, Electron / Q000379:methods*; D057054:Molecular Imaging / Q000379:methods; D053758:Nanoparticles / Q000737:chemistry* / Q000648:ultrastructure*; D010316:Particle Size; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D006046:Gold
keywords: electron microscopy; endocytosis inhibition; laser scanning microscopy; nanoparticle uptake; polymer-coated gold nanoparticle; stereology