Quantification of silver nanoparticle uptake and distribution within individual human macrophages by FIB/SEM slice and view

Abstract

BackgroundQuantification of nanoparticle (NP) uptake in cells or tissues is very important for safety assessment. Often, electron microscopy based approaches are used for this purpose, which allow imaging at very high resolution. However, precise quantification of NP numbers in cells and tissues remains challenging. The aim of this study was to present a novel approach, that combines precise quantification of NPs in individual cells together with high resolution imaging of their intracellular distribution based on focused ion beam/ scanning electron microscopy (FIB/SEM) slice and view approaches.ResultsWe quantified cellular uptake of 75 nm diameter citrate stabilized silver NPs (Ag 75 Cit) into an individual human macrophage derived from monocytic THP-1 cells using a FIB/SEM slice and view approach. Cells were treated with 10 μg/ml for 24 h. We investigated a single cell and found in total 3138 ± 722 silver NPs inside this cell. Most of the silver NPs were located in large agglomerates, only a few were found in clusters of fewer than five NPs. Furthermore, we cross-checked our results by using inductively coupled plasma mass spectrometry and could confirm the FIB/SEM results.ConclusionsOur approach based on FIB/SEM slice and view is currently the only one that allows the quantification of the absolute dose of silver NPs in individual cells and at the same time to assess their intracellular distribution at high resolution. We therefore propose to use FIB/SEM slice and view to systematically analyse the cellular uptake of various NPs as a function of size, concentration and incubation time.

Quantification of silver nanoparticles taken up by single cells using inductively coupled plasma mass spectrometry in the single cell measurement mode

“…However, mass cytometry is not able to discriminate between AgNPs externally adsorbed versus AgNPs taken up and these limitations might be circumvented by using the proling method step FIB/SEM slice and view. 10 It is noteworthy that the methodology proposed in this study, SC-ICP-MS, is not able to elucidate the AgNP chemical composition inside the cells as well as a possible transformation by oxidation to ionic Ag. Aer AgNP uptake, cells might contain a fraction of ionic silver due to AgNP oxidation that would be detected by SC-ICP-MS as a discrete spike signal with an intensity proportional to the total amount of Ag atoms (from ionic silver or/and AgNPs) contained per individual cell.…”

“…2e-h show an appearing right-hand tail, indicating that some individual cells contained multiple AgNPs, most likely aggregates as it was stated previously by Guehrs et al in 2017. 10 This right-hand tail was more pronounced for THP-1 monocytes treated at the highest exposure dose and time (1 mg mL À1 , 24 h) ( Fig. 2h).…”

“…The results obtained by SC-ICP-MS were compared and validated by parallel measurements with ICP-MS aer digestion of the cell suspension as well as with other alternative promising techniques, providing single cell analysis such as mass cytometry, CyTOF, and the proling method step FIB/SEM slice and view. 10…”

Interaction of γ-Fe2O3 nanoparticles with Citrus maxima leaves and the corresponding physiological effects via foliar application

“…Nowadays, nanomaterials become a hotspot of research interests and attract the attention of many researchers. A variety of nanoparticles (NPs) have been studied on human cells [4, 5], animal cells [6] and plants [7] about their toxicity or applications. As one of the most widely explored and applied nanomaterials, iron oxide nanoparticles (γ-Fe 2 O 3 NPs) are widely used in medical diagnostics, controlled drug release, separation technologies and environmental engineering [8].…”

Review of emerging concepts in nanotoxicology: opportunities and challenges for safer nanomaterial design

“…The silver nanoparticles are a high surface area source for the silver ions ( Figure 4(J-K)). In biological assays, silver NPs showed a continuous release of silver ions during transport to their final location (Singh et al 2012;Guehrs et al 2017). Nanotoxicological assays are able to give information on the fraction of ions to NPs in the supernatant or permeate from the biological medium supplemented silver NPs using ICP or mass spectrometry (Bohmert et al 2017).…”