Upconverting nanoparticles (UCNPs) are of particular interest in nanomedicine for in vivo deep-tissue optical cancer bioimaging due to their efficient cellular uptake dependent on polymer coating. In this study, particles, ca. 25 nm in diameter, were prepared by a high-temperature coprecipitation of lanthanide chlorides. To ensure optimal dispersion of UCNPs in aqueous milieu, they were coated with three different polymers containing reactive groups, i.e., poly(ethylene glycol)-alendronate (PEG-Ale), poly(N,N-dimethylacrylamide-co-2-aminoethylacrylamide)-alendronate (PDMA-Ale), and poly(methyl vinyl ether-co-maleic acid) (PMVEMA). All the particles were characterized by TEM, DLS, FTIR, and spectrofluorometer to determine the morphology, hydrodynamic size and ξ-potential, composition, and upconversion luminescence. The degradability/dissolution of UCNPs in water, PBS, DMEM, or artificial lysosomal fluid (ALF) was evaluated using an ion-selective electrochemical method and UV-Vis spectroscopy. The dissolution that was more pronounced in PBS at elevated temperatures was decelerated by polymer coatings. The dissolution in DMEM was relatively small, but much more pronounced in ALF. PMVEMA with multiple anchoring groups provided better protection against particle dissolution in PBS than PEG-Ale and PDMA-Ale polymers containing only one reactive group. However, the cytotoxicity of the particles depended not only on their ability to rapidly degrade, but also on the type of coating. According to MTT, neat UCNPs and UCNP@PMVEMA were toxic for both rat cells (C6) and rat mesenchymal stem cells (rMSCs), which was in contrast to the UCNP@Ale-PDMA particles that were biocompatible. On the other hand, both the cytotoxicity and uptake of the UCNP@Ale-PEG particles by C6 and rMSCs were low, according to MTT assay and ICP-MS, respectively. This was confirmed by a confocal microscopy, where the neat UCNPs were preferentially internalized by both cell types, followed by the UCNP@PMVEMA, UCNP@Ale-PDMA, and UCNP@Ale-PEG particles. This study provides guidance for the selection of a suitable nanoparticle coating with respect to future biomedical applications where specific behaviors (extracellular deposition vs. cell internalization) are expected.

• Klíčová slova
• degradation, lanthanides, luminescence, nanoparticles, upconversion,
• MeSH
• alendronát MeSH
• krysa rodu Rattus MeSH
• nanočástice * chemie   MeSH
• polyethylenglykoly chemie   MeSH
• polymery * chemie   MeSH
• voda MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• alendronát MeSH
• polyethylenglykoly MeSH
• polymery * MeSH
• voda MeSH

The detection of cancer biomarkers in histological samples and blood is of paramount importance for clinical diagnosis. Current methods are limited in terms of sensitivity, hindering early detection of disease. We have overcome the shortcomings of currently available staining and fluorescence labeling methods by taking an integrative approach to establish photon-upconversion nanoparticles (UCNP) as a powerful platform for cancer detection. These nanoparticles are readily synthesized in different sizes to yield efficient and tunable short-wavelength light emission under near-infrared excitation, which eliminates optical background interference of the specimen. Here we present a protocol for the synthesis of UCNPs by high-temperature co-precipitation or seed-mediated growth by thermal decomposition, surface modification by silica or poly(ethylene glycol) that renders the particles resistant to nonspecific binding, and the conjugation of streptavidin or antibodies for biological detection. To detect blood-based biomarkers, we present an upconversion-linked immunosorbent assay for the analog and digital detection of the cancer marker prostate-specific antigen. When applied to immunocytochemistry analysis, UCNPs enable the detection of the breast cancer marker human epidermal growth factor receptor 2 with a signal-to-background ratio 50-fold higher than conventional fluorescent labels. UCNP synthesis takes 4.5 d, the preparation of the antibody-silica-UCNP conjugate takes 3 d, the streptavidin-poly(ethylene glycol)-UCNP conjugate takes 2-3 weeks, upconversion-linked immunosorbent assay takes 2-4 d and immunocytochemistry takes 8-10 h. The procedures can be performed after standard laboratory training in nanomaterials research.

• MeSH
• imunosorbenty MeSH
• lidé MeSH
• nádorové biomarkery MeSH
• nádory * diagnóza   MeSH
• nanočástice * chemie   MeSH
• oxid křemičitý chemie   MeSH
• polyethylenglykoly chemie   MeSH
• streptavidin MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• imunosorbenty MeSH
• nádorové biomarkery MeSH
• oxid křemičitý MeSH
• polyethylenglykoly MeSH
• streptavidin MeSH

Lanthanide-doped upconversion nanoparticles (UCNPs) have a unique capability of upconverting near-infrared (NIR) excitation into ultraviolet, visible, and NIR emission. Conventional UCNPs composed of NaYF4:Yb3+/Er3+(Tm3+) are excited by NIR light at 980 nm, where undesirable absorption by water can cause overheating or damage of living tissues and reduce nanoparticle luminescence. Incorporation of Nd3+ ions into the UCNP lattice shifts the excitation wavelength to 808 nm, where absorption of water is minimal. Herein, core-shell NaYF4:Yb3+/Er3+@NaYF4:Nd3+ nanoparticles, which are doubly doped by sensitizers (Yb3+ and Nd3+) and an activator (Er3+) in the host NaYF4 matrix, were synthesized by high-temperature coprecipitation of lanthanide chlorides in the presence of oleic acid as a stabilizer. Uniform core (24 nm) and core-shell particles with tunable shell thickness (~0.5-4 nm) were thoroughly characterized by transmission electron microscopy (TEM), energy-dispersive analysis, selected area electron diffraction, and photoluminescence emission spectra at 808 and 980 nm excitation. To ensure dispersibility of the particles in biologically relevant media, they were coated by in-house synthesized poly(ethylene glycol) (PEG)-neridronate terminated with an alkyne (Alk). The stability of the NaYF4:Yb3+/Er3+@NaYF4:Nd3+-PEG-Alk nanoparticles in water or 0.01 M PBS and the presence of PEG on the surface were determined by dynamic light scattering, ζ-potential measurements, thermogravimetric analysis, and FTIR spectroscopy. Finally, the adhesive azidopentanoyl-modified GGGRGDSGGGY-NH2 (RGDS) peptide was immobilized on the NaYF4:Yb3+/Er3+@NaYF4:Nd3+-PEG-Alk particles via Cu(I)-catalyzed azide-alkyne cycloaddition. The toxicity of the unmodified core-shell NaYF4:Yb3+/Er3+@NaYF4:Nd3+, NaYF4:Yb3+/Er3+@NaYF4:Nd3+-PEG-Alk, and NaYF4:Yb3+/Er3+@NaYF4:Nd3+-PEG-RGDS nanoparticles on both Hep-G2 and HeLa cells was determined, confirming no adverse effect on their survival and proliferation. The interaction of the nanoparticles with Hep-G2 cells was monitored by confocal microscopy at both 808 and 980 nm excitation. The NaYF4:Yb3+/Er3+@NaYF4:Nd3+-PEG-RGDS nanoparticles were localized on the cell membranes due to specific binding of the RGDS peptide to integrins, in contrast to the NaYF4:Yb3+/Er3+@NaYF4:Nd3+-PEG-Alk particles, which were not engulfed by the cells. The NaYF4:Yb3+/Er3+@NaYF4:Nd3+-PEG-RGDS nanoparticles thus appear to be promising as a new non-invasive probe for specific bioimaging of cells and tissues. This development makes the nanoparticles useful for diagnostic and/or, after immobilization of a bioactive compound, even theranostic applications in the treatment of various fatal diseases.

• Klíčová slova
• 808 nm excitation, Hep-G2 and HeLa cells, PEG-neridronate, RGDS peptide, core-shell, luminescence, upconversion nanoparticles,
• Publikační typ
• časopisecké články MeSH