Effect of Excess Ligand on the Reverse Microemulsion Silica Coating of NaLnF4 Nanoparticles.

Silica coating of inorganic nanoparticles (NPs) is widely employed as a means of providing colloidal stability in aqueous media and surface functionality for a variety of applications, particularly in biology. When the NPs are synthesized with a surface coating of an organic surfactant like oleic acid, silica coating is performed by using the reverse microemulsion method. There are many reports in the literature of the successful application of this method to NaYF4 upconversion NPs (doped with Yb and Er), and we have used this method to coat NaHoF4 NPs designed as a mass cytometry reagent. This method failed when we attempted to apply it to other NaLnF4 NPs (Ln = Sm, Eu, Tb). In this report we describe an investigation of the problem and show how it can be overcome. To control size in the synthesis of NaLnF4 NPs and at the same time maintain size uniformity, it is necessary to adjust the Na/F and F/Ln ratios. Problems with silica coating are associated with substoichiometric F/Ln ratios (F/Ln < 4) that leave Ln oleate salts as a byproduct, often as a phase-separated oily layer that could not be purified from the NPs by precipitation with ethanol and redispersion in hexanes. The nature of the oily byproduct was inferred from a combination of TGA, NMR, and FTIR measurements. We explored five different additional purification procedures, and by adopting the appropriate purification method, NaLnF4 NPs with a variety of compositions and synthesized using different reaction conditions could be coated with a thin shell of silica.

An effective polymer cross-linking strategy to obtain stable dispersions of upconverting NaYF4 nanoparticles in buffers and biological growth media for biolabeling applications.

Ligands on the nanoparticle surface provide steric stabilization, resulting in good dispersion stability. However, because of their highly dynamic nature, they can be replaced irreversibly in buffers and biological medium, leading to poor colloidal stability. To overcome this, we report a simple and effective cross-linking methodology to transfer oleate-stabilized upconverting NaYF(4) core/shell nanoparticles (UCNPs) from hydrophobic to aqueous phase, with long-term dispersion stability in buffers and biological medium. Amphiphilic poly(maleic anhydride-alt-1-octadecene) (PMAO) modified with and without poly(ethylene glycol) (PEG) was used to intercalate with the surface oleates, enabling the transfer of the UCNPs to water. The PMAO units on the phase transferred UCNPs were then successfully cross-linked using bis(hexamethylene)triamine (BHMT). The primary advantage of cross-linking of PMAO by BHMT is that it improves the stability of the UCNPs in water, physiological saline buffers, and biological growth media and in a wide range of pH values when compared to un-cross-linked PMAO. The cross-linked PMAO-BHMT coated UCNPs were found to be stable in water for more than 2 months and in physiological saline buffers for weeks, substantiating the effectiveness of cross-linking in providing high dispersion stability. The PMAO-BHMT cross-linked UCNPs were extensively characterized using various techniques providing supporting evidence for the cross-linking process. These UCNPs were found to be stable in serum supplemented growth medium (37 °C) for more than 2 days. Utilizing this, we demonstrate the uptake of cross-linked UCNPs by LNCaP cells (human prostate cancer cell line), showing their utility as biolabels.

Lanthanide nanoparticles for high sensitivity multiparameter single cell analysis.

Mass cytometry (MC) is a high throughput multiparameter analytical technique for determining biomarker expression in cells. In MC, antibodies (Abs) are tagged with heavy metal isotopes via conjugation to metal chelating polymers (MCPs). To improve the sensitivity of MC towards low abundance biomarkers, we are developing nanoparticle (NP)-based reagents as mass tags for Abs. We examine the use of silica-coated NaHoF4 NPs (d ∼ 12 nm) decorated with PEG5k conjugated to thiol-modified primary or secondary Abs for MC assays. We compare the sensitivity of NP-Ab conjugates to MCP-Ab conjugates towards seven biomarkers with varying expression levels across six cell lines. We also perform a multi-parameter assay using a cocktail of both NP- and MCP-based reagents to detect seven cellular markers in peripheral blood mononuclear cells (PBMCs). In the case of highly abundant markers, signal enhancements from NP-Ab conjugates offer minimal advantages over MCP-Ab conjugates, which already give strong signals. In the case of biomarkers with lower abundance, the level of signal enhancements depended on the nature of the biomarker being detected, or on the type of detection method used. When comparing the indirect detection of CD14 on THP-1 cells using NPs or MCPs conjugated to secondary Abs, the NP reagents offered little signal enhancements compared to the MCP reagents. However, in the case of direct CD14 detection on THP-1 or U937 cells using NPs or MCPs conjugated to primary Abs, a 30- or 450-fold signal enhancement was seen from the NP-based reagent. In the experiments where both NP-Ab and MCP-Ab conjugates were used together to stain PBMCs, we found that the presence of the NP-Ab conjugates did not affect the function of MCP-Ab conjugates, and the NP-Ab conjugates showed minimal non-specific interaction with cells without the target biomarker (CD14). Furthermore, these NP-Ab conjugates could be used to identify rare CD14+ monocytes from the PBMC mixture with a 20-fold signal increase when compared to the use of only MCP-Ab conjugates. Collectively, the strong signal amplification obtained from NP reagents demonstrate the potential of these reagents to be used in conjunction with MCP-reagents to detect rare cellular markers or cell types that may otherwise be overlooked when using MCP-reagents alone.

Polarization-dependent extraordinary optical transmission from upconversion nanoparticles.

Enhanced upconversion (UC) emission was experimentally demonstrated using gold double antenna nanoparticles coupled to nanoslits in gold films. The transmitted red emission from UC ytterbium and erbium co-doped sodium yttrium fluoride (NaYF4:Yb(3+)/Er(3+)) nanoparticles (UC NPs) at ∼665 nm (excited with a 980 nm diode laser) was enhanced relative to the green emission at ∼550 nm. The relatively enhanced UC NP emission could be tuned by the different polarization-dependent extraordinary optical transmission modes coupled to the gold nanostructures. Finite-difference time-domain calculations suggest that the preferential enhanced UC emission is related to a combination of different surface plasmon mode excitation coupling to cavity Fabry-Perot interactions. A maximum UC enhancement of 6-fold was measured for nanoslit arrays in the absence of the double antennas. In the presence of the double nanoantennas inside the nanoslits, the UC enhancement was between 2- and 4-fold, depending on the experimental conditions.

Ln(3+)-doped nanoparticles for upconversion and magnetic resonance imaging: some critical notes on recent progress and some aspects to be considered.

In this feature article we will critically discuss the synthesis and characterisation aspects of Ln(3+)-doped nanoparticles (NPs) that show upconversion, upon 980 nm excitation. Upconversion is a non-linear process that converts two or more low-energy photons, often near-infrared photons, into one of higher energy, e.g. blue and 800 nm from Tm(3+) and green and red from Er(3+) or Ho(3+). Nearly all researchers use the absorption of 980 nm light by Yb(3+) as the sensitiser for the co-doped emissive Ln(3+) ions. The focus will be on LnF(3) and MLnF(4) (M = alkali metal) as the host matrix, because most progress has been made with these. In particular we will argue that a detailed understanding of how the dopant ions and the host Ln(3+) ions are distributed (in the core) and how (doped) shell growth occurs is not well understood. Moreover, their use as optical and magnetic resonance imaging contrast agents will be discussed. We will argue that deep-tissue imaging beyond 600 µm with retention of optical resolution, i.e. to see fine structure such as blood capillaries in brain tissues, has not yet been achieved. Three key parameters have been identified as impediments: (i) the low absorption efficiency of the Yb(3+) sensitiser, (ii) the low quantum yield of upconversion, and (iii) the long-lived excited states. On the other hand, there are very encouraging results that suggest that these nanoparticles could be developed into very potent magnetic resonance imaging (MRI) contrast agents.

The unexpected structures of "core-shell" and "alloy" LnF3 nanoparticles as examined by variable energy X-ray photo-electron spectroscopy.

Lanthanide fluoride nanoparticles were synthesized in aqueous media using procedures intended for a core-shell structure of Ln((1))F(3)-Ln((2))F(3), its reverse architecture, and an alloy structure. Their structures were examined by variable photon energy photo-electron spectroscopy using synchrotron radiation, along with X-ray powder diffractometry, transmission electron microscopy, energy dispersive X-ray spectroscopy, and luminescence spectroscopy. The results show that the nanoparticles intended for a core-shell structure do not have a core-shell structure, and that nanoparticles intended for an alloy structure do not always have an alloy structure. A possible explanation for this is cation exchange, a phenomenon that occurs when LnF(3) nanoparticles are exposed to another Ln(3+) ion in aqueous media, resulting in Ln(3+) ions in nanoparticles being quickly replaced by Ln(3+) ions in solution. This cation exchange effectively competes with the precipitation of LnF(3), which leads to a concentration gradient in the case of the combination of LaF(3) and GdF(3), and to nearly an alloy structure (isotropic mixture of all the ions) in the case of the combination of LaF(3) and NdF(3), regardless of the procedure used. Finally, the intended "core-shell" nanoparticles were doped with Eu(3+) to show that a non-core-shell structure can also give rise to the improvement of optical properties as compared with the corresponding core nanoparticles. These results suggest that conclusions in the literature that a core-shell structure was obtained as inferred by TEM or enhanced luminescence may not be correct.