Chemie und Prozesstechnik
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- 2019 (2) (entfernen)
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- ICP-MS (2)
- Laser ablation (2)
- Multicellular spheroid (2)
- Cell (1)
- Nanoparticle (1)
- Nanoparticles (1)
Organisationseinheit der BAM
- 1.1 Anorganische Spurenanalytik (2) (entfernen)
Metallic nanoparticles (NPs) are currently applied in a variety of consumer products and are also attractive for medical applications. With their widespread use, the potential for human exposure to NPs — either intended or unintended — is increasing. Therefore, many studies have evaluated the toxicity and transport mechanism of NPs. In comparison with two-dimensional cultured cells, multicellular spheroids (MCS) look promising to be used as a three-dimensional cellular model, having unique advantages in nanoparticle studies due to the fact that interactions with excreted extracellular matrix can be investigated. Fibroblast cells are one of the most important cell systems to express a microenvironment by excreting an abundant extracellular matrix. For bioimaging laser ablation inductively coupled-plasma mass spectrometry (LA-ICP-MS) is used in this investigation to study the interaction of metallic NPs with MCS for multi-element detection offering a wide dynamic range. As a mass spectrometer we have applied a time-of-flight (TOF) instrument for (quasi-) simultaneous detection of all isotopes of elements of interest.
The aim of this study is to investigate the localization of silver (Ag) NPs exposed to fibroblast MCSs by means of LA-ICP-TOF-MS. In addition, for demonstrating elemental microscopy we chose phosphorous (31P) and iron (56Fe) to visualize regions of enriched extracellular matrix and single cells, respectively. In this presentation, we show that exposed Ag NPs are highly accumulated at the same position of single cells in an outer rim of fibroblast MCSs.
We have applied laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) with subcellular resolution as an elemental mass microscope to investigate the distributions of Ag nanoparticles (NP) in a 3-dimentional multicellular spheroid (MCS) model. The production of MCS has been optimized by changing the seeding cell number (500 to 40,000 cells) and the growth period (1 to 10 days). Incubations of MCS with Ag nanoparticle suspensions were performed with a concentration of 5 µg mL-1 for 24 hours. Thin-sections of the Eosin stained MCS were analysed by elemental mass microscopy using LA-ICP-MS to image distributions of 109Ag, 31P, 63Cu, 66Zn and 79Br. A calibration using NP suspensions was applied to convert the measured Ag intensity into the number of particles being present in each measurement pixel. The numbers of NP determined ranged from 30 up to 4,000 particles in an enrichment zone. The particle distribution was clearly correlated to 31P, 66Zn and 79Br and was localized in an outer rim of proliferating cells (confirmed by DAPI) with a width of about two-single cell diameters. For the highest seeding cell number NPs were only detected in this outer rim, whereas small molecules as for instance 79Br and 109Ag ions were detected in the core of the MCS as well. Aniline blue staining demonstrated that this outer rim was rich in collagen structures in which fibroblast cells were embedded and a thin-membrane was visible which separated the core from the biological active cell layer functioning as biological barriers for NP transport. In this presentation, we will show the possibility using this 3-dimensional model for toxicological and medical applications.