Quantitative Imaging of Silver Nanoparticles and Essential Elements in Thin Sections of Fibroblast Multicellular Spheroids by High Resolution Laser Ablation Inductively Coupled Plasma Time-of-Flight Mass Spectrometry

Abstract

We applied high resolution laser ablation inductively coupled plasma time-of-flight mass spectrometry (LA-ICP-TOF-MS) with cellular spatial resolution for bioimaging of nanoparticles uptaken by fibroblast multicellular spheroids (MCS). This was used to quantitatively investigate interactions of silver nanoparticles (Ag NPs) and the distributions of intrinsic minerals and biologically relevant elements within thin sections of a fibroblast MCS as a three-dimensional in vitro tissue model. We designed matrix-matched calibration standards for this purpose and printed them using a noncontact piezo-driven array spotter with a Ag NP suspension and multielement standards. The limits of detection for Ag, Mg, P, K, Mn, Fe, Co, Cu, and Zn were at the femtogram (10–15 g) level, which is sufficient to investigate intrinsic minerals in thin MCS sections (20 μm thick). After incubation for 48 h, Ag NPs were enriched in the outer rim of the MCS but not detected in the core. The localization of Ag NPs was inhomogeneous in the outer rim, and they were colocalized with a single-cell-like structure visualized by Fe distribution (pixel size of elemental images: 5 × 0.5 μm). The quantitative value for the total mass of Ag NPs in a thin section by the present method agreed with that obtained by ICP-sector field (SF)-MS with a liquid mode after acid digestion.

Micro-droplet-based calibration for quantitative elemental bioimaging by LA-ICPMS

“…Alternatively, micro-array spotters have been used to produce calibration standards in an automated and reproducible way. ICP-MS standard solutions were printed on nitrocellulose membranes [ 23 ] and on target glass slides [ 24 ] with volumes in the picoliter range [ 23 , 25 ]. In this regard, single-cell analysis by LA-ICPMS was performed to quantify an Ir-DNA intercalator and an mDOTA-Ho dye in adherent 3T3 fibroblast cells [ 23 ].…”

“…In this regard, single-cell analysis by LA-ICPMS was performed to quantify an Ir-DNA intercalator and an mDOTA-Ho dye in adherent 3T3 fibroblast cells [ 23 ]. Calibration standards using Ag NP suspensions were prepared in a similar manner to investigate the quantitative distribution of Ag NPs in a multicellular tumor spheroid model [ 25 ].…”

Head-to-Head Comparison of the Penetration Efficiency of Lipid-Based Nanoparticles into Tumor Spheroids

“… 26 The majority of studies of NP distributions in tumor spheroids use tumor spheroid fixation, 27 − 35 often in combination with histological sectioning. 7 , 36 − 39 Nevertheless, there are also reports of NP penetration in live tumor spheroids using confocal-scanning microscopy. 40 − 50 Live imaging of NP-treated tumor spheroids ensures that the studied NPs preserve localization, stability, and emission wavelength, 51 , 52 as indicated for semiconducting NPs.…”

Head-to-head comparison of the penetration efficiency of lipid-based nanoparticles in a 3D tumor spheroid model

“…rapid proliferation at the surface and slow metabolism -or necrosis -in the center of spheroids 25 . The majority of studies of NP distributions in tumor spheroids use tumor spheroid fixation [26][27][28][29][30][31][32][33][34] , often in combination with histological sectioning 7,[35][36][37][38] . Nevertheless, there are also reports of NP penetration in live tumor spheroids using confocal-scanning microscopy [39][40][41][42][43][44][45][46][47][48][49] .…”