Abstract:Differentiation between cell surface-bound and internalized nanoparticles is challenging yet essential for accurately quantifying cellular uptake. Here, we describe a versatile mass spectrometry-based method that allows separate quantification of both cell surface-bound and internalized nanoparticles. This rapid method uses tuned laser fluencies to selectively desorb and ionize cell surface-bound cationic gold nanoparticles from intact cells, providing quantification of external particles. Overall nanoparticle… Show more
“…The data demonstrate that with rising NP concentrations the excretion via exocytosis is also increasing, which takes place largely within the first 24 h. Note that ICP-MS can not distinguish between Au NPs just adherent to the outer cell membrane and Au NPs that have been in fact endocytosed. There are methods available which allow for separating both populations [32, 33]. However, we did not apply this analysis, as it would not be relevant for the in vivo studies, as discussed in the respective paragraph.Fig.…”
BackgroundMesenchymal stromal cells (MSCs) have an inherent migratory capacity towards tumor tissue in vivo. With the future objective to quantify the tumor homing efficacy of MSCs, as first step in this direction we investigated the use of inorganic nanoparticles (NPs), in particular ca. 4 nm-sized Au NPs, for MSC labeling. Time dependent uptake efficiencies of NPs at different exposure concentrations and times were determined via inductively coupled plasma mass spectrometry (ICP-MS).ResultsThe labeling efficiency of the MSCs was determined in terms of the amount of exocytosed NPs versus the amount of initially endocytosed NPs, demonstrating that at high concentrations the internalized Au NPs were exocytosed over time, leading to continuous exhaustion. While exposure to NPs did not significantly impair cell viability or expression of surface markers, even at high dose levels, MSCs were significantly affected in their proliferation and migration potential. These results demonstrate that proliferation or migration assays are more suitable to evaluate whether labeling of MSCs with certain amounts of NPs exerts distress on cells. However, despite optimized conditions the labeling efficiency varied considerably in MSC lots from different donors, indicating cell specific loading capacities for NPs. Finally, we determined the detection limits of Au NP-labeled MSCs within murine tissue employing ICP-MS and demonstrate the distribution and homing of NP labeled MSCs in vivo.ConclusionAlthough large amounts of NPs improve contrast for imaging, duration and extend of labeling needs to be adjusted carefully to avoid functional deficits in MSCs. We established an optimized labeling strategy for human MSCs with Au NPs that preserves their migratory capacity in vivo.Electronic supplementary materialThe online version of this article (doi:10.1186/s12951-017-0258-5) contains supplementary material, which is available to authorized users.
“…The data demonstrate that with rising NP concentrations the excretion via exocytosis is also increasing, which takes place largely within the first 24 h. Note that ICP-MS can not distinguish between Au NPs just adherent to the outer cell membrane and Au NPs that have been in fact endocytosed. There are methods available which allow for separating both populations [32, 33]. However, we did not apply this analysis, as it would not be relevant for the in vivo studies, as discussed in the respective paragraph.Fig.…”
BackgroundMesenchymal stromal cells (MSCs) have an inherent migratory capacity towards tumor tissue in vivo. With the future objective to quantify the tumor homing efficacy of MSCs, as first step in this direction we investigated the use of inorganic nanoparticles (NPs), in particular ca. 4 nm-sized Au NPs, for MSC labeling. Time dependent uptake efficiencies of NPs at different exposure concentrations and times were determined via inductively coupled plasma mass spectrometry (ICP-MS).ResultsThe labeling efficiency of the MSCs was determined in terms of the amount of exocytosed NPs versus the amount of initially endocytosed NPs, demonstrating that at high concentrations the internalized Au NPs were exocytosed over time, leading to continuous exhaustion. While exposure to NPs did not significantly impair cell viability or expression of surface markers, even at high dose levels, MSCs were significantly affected in their proliferation and migration potential. These results demonstrate that proliferation or migration assays are more suitable to evaluate whether labeling of MSCs with certain amounts of NPs exerts distress on cells. However, despite optimized conditions the labeling efficiency varied considerably in MSC lots from different donors, indicating cell specific loading capacities for NPs. Finally, we determined the detection limits of Au NP-labeled MSCs within murine tissue employing ICP-MS and demonstrate the distribution and homing of NP labeled MSCs in vivo.ConclusionAlthough large amounts of NPs improve contrast for imaging, duration and extend of labeling needs to be adjusted carefully to avoid functional deficits in MSCs. We established an optimized labeling strategy for human MSCs with Au NPs that preserves their migratory capacity in vivo.Electronic supplementary materialThe online version of this article (doi:10.1186/s12951-017-0258-5) contains supplementary material, which is available to authorized users.
“…When referring to cells, this could involve the percentage of cells which have internalized particles, which is, for example, frequently used in flow cytometry studies, or the percentage of cells which have a certain number of particles incorporated, which is often used in case the number of discrete particles in a cell can be directly counted . When referring to particles, this could be the percentage of particles which have been internalized, which, for example, is done when the amount of particles remaining in the cell medium and the amount of particles inside cells are quantified with elemental analysis . Second, also the t ‐axis in Figure , the second quantifier, addressing the question “how fast”, needs to be defined.…”
Section: Useful Parameters For the Quantification Of Particle Uptakementioning
There is a large number of two‐dimensional static in vitro studies about the uptake of colloidal nano‐ and microparticles, which has been published in the last decade. In this Minireview, different methods used for such studies are summarized and critically discussed. Supplementary experimental data allow for a direct comparison of the different techniques. Emphasis is given on how quantitative parameters can be extracted from studies in which different experimental techniques have been used, with the goal of allowing better comparison.
“…Diese Methode wird bspw. genutzt, wenn die Menge an übergebliebenen Partikeln im Zellmedium und die Menge an internalisierten Partikeln über elementare Analysen quantifiziert wurde . Zweitens muss auch der zweite quantisierende Parameter, die t ‐Achse in Abbildung mit der Frage “wie schnell”, definiert werden.…”
Section: Nützliche Parameter Für Die Quantifizierung Der Partikelaufnunclassified
Im letzten Jahrzehnt gab es eine große Anzahl publizierter zweidimensionaler und zeitunabhängiger In‐vitro‐Studien über die Aufnahme kolloidaler Nano‐ und Mikropartikel in Zellen. In diesem Kurzaufsatz werden unterschiedliche angewandte Methoden solcher Untersuchungen zusammengefasst und kritisch diskutiert. Ergänzende experimentelle Daten erlauben einen direkten Vergleich zwischen diesen angewandten Techniken. Das Hauptaugenmerk wird darauf liegen, quantitative Parameter aus solchen Studien zu gewinnen, in denen mehrere verschiedene experimentelle Messmethoden genutzt wurden, mit dem Ziel eine bessere Vergleichbarkeit der Daten zu gewährleisten.
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