title: Comparison of fluorescence-based methods to determine nanoparticle uptake by phagocytes and non-phagocytic cells in vitro.
abstract: At many portals of entry the relative uptake by phagocytes and non-phagocytic cells has a prominent effect on availability and biological action of nanoparticles (NPs). Cellular uptake can be determined for fluorescence-labeled NPs. The present study compares three methods (plate reader, flow cytometry and image analysis) in order to investigate the influence of particle size and functionalization and medium content on cellular uptake of fluorescence-labeled polystyrene particles and to study the respective method́s suitability for uptake studies. For comparison between the techniques, ratios of macrophage to alveolar epithelial cell uptakes were used. Presence of serum protein in the exposure solution decreased uptake of carboxyl-functionalized and non-functionalized particles; there was no clear effect for the amine-functionalized particles. The 200nm non- or carboxyl-functionalized NPs were taken up preferentially by phagocytes while for amine-functionalized particles preference was lowest. The presence of the serum slightly increased the preference for these particles. In conclusion, due to the possibility of calibration, plate reader measurements might present a better option than the other techniques to (semi)quantify differences between phagocytes and non-phagocytic cells for particles with different fluorescence. In order to obtain unbiased data the fluorescent labeling has to fulfill certain requirements.
authors.affiliation: Center for Medical Research, Medical University of Graz, Stiftingtalstr. 24, 8010 Graz, Austria. Electronic address: Claudia.meindl@klinikum-graz.at.
authors.forename: Meindl
authors.initials: M
authors.lastname: Claudia
authors.affiliation: Center for Medical Research, Medical University of Graz, Stiftingtalstr. 24, 8010 Graz, Austria. Electronic address: kristin.oehlinger@medunigraz.at.
authors.forename: Öhlinger
authors.initials: Ö
authors.lastname: Kristin
authors.affiliation: Center for Medical Research, Medical University of Graz, Stiftingtalstr. 24, 8010 Graz, Austria. Electronic address: Jennifer.ober@medunigraz.at.
authors.forename: Ober
authors.initials: O
authors.lastname: Jennifer
authors.affiliation: Institute of Pharmaceutical Sciences, Department of Pharmaceutical Technology, Karl-Franzens-University of Graz, Humboldtstr, 46, 8010 Graz, Austria. Electronic address: eva.roblegg@uni-graz.at.
authors.forename: Roblegg
authors.initials: R
authors.lastname: Eva
authors.affiliation: Center for Medical Research, Medical University of Graz, Stiftingtalstr. 24, 8010 Graz, Austria. Electronic address: eleonore.froehlich@medunigraz.at.
authors.forename: Fröhlich
authors.initials: F
authors.lastname: Eleonore
mesh_terms: D000072283:A549 Cells; D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D018920:Coculture Techniques; D004705:Endocytosis; D004847:Epithelial Cells / Q000502:physiology*; D005434:Flow Cytometry; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000494:pharmacology; D051379:Mice; D018613:Microscopy, Confocal; D053758:Nanoparticles / Q000737:chemistry; D010316:Particle Size; D010586:Phagocytes / Q000502:physiology*; D011137:Polystyrenes / Q000737:chemistry / Q000494:pharmacology
chemical_list: D005456:Fluorescent Dyes; D011137:Polystyrenes
keywords: Cellular uptake; Exposure conditions; Inhalation exposure; Nanoparticles; Phagocytes