Single molecule localization based optical super-resolution microscopy (SRM) techniques, and in particular stochastic optical reconstruction microscopy (STORM), are powerful imaging tools to resolve structures below the diffraction limit
In contrast to small molar mass compounds, detailed structural investigations of inorganic coreorganic ligand shell hybrid nanoparticles remain challenging. Assessment of batch reaction induced heterogeneities of surface chemical properties and their correlation with particle size has been a particularly long-standing issue. Applying a combination of high performance liquid chromatography (HPLC) and gel permeation chromatography (GPC) to ultrasmall (< 10 nm diameter) poly(ethylene glycol) coated (PEGylated) fluorescent core-shell silica nanoparticles, here we elucidate previously unknown surface heterogeneities resulting from varying dye conjugation to nanoparticle silica cores and surfaces. Heterogeneities are predominantly governed by dye charge as corroborated by molecular dynamics simulations. We demonstrate that this insight enables development of synthesis protocols to achieve PEGylated and targeting ligand functionalized PEGylated silica nanoparticles with dramatically improved surface chemical homogeneity as evidenced by single peak HPLC chromatograms. Since surface chemical properties are key to all nanoparticle interactions, we expect these methods and fundamental insights to become relevant to a number of systems for applications including bioimaging and nanomedicine.
Synthetic advances in the formation of ultrasmall (<10 nm) fluorescent poly(ethylene glycol)-coated (PEGylated) core-shell silica nanoparticles (SNPs), enabling improved particle size and surface chemical property control have led to successful clinical translation of SNPs as diagnostic probes in oncology. Despite the success of such probes, details of the dye incorporation and resulting silica architecture are still poorly understood. Here, we employ afterpulse-corrected fluorescence correlation spectroscopy (FCS) to monitor fast fluorescence fluctuations (lag times <10−5 s) of the negatively charged cyanine dye Cy5 as a probe to study such details for dye encapsulation in 5 nm silica cores of PEGylated core-shell SNPs (C dots). Upon deposition of additional silica shells over the silica core we find that the amplitude of photo-induced cis-trans isomerization decreases, suggesting that the Cy5 dyes are located near or on the surface of the original SNP cores. In combination with time correlated fluorescence decay measurements we deduce radiative and non-radiative rates of the Cy5 dye in these particles. Results demonstrate that FCS is a well-suited tool to investigate aspects of the photophysics of fluorescent nanoparticles, and that conformational changes of cyanine dyes like Cy5 are excellent indicators for the local dye environment within ultrasmall SNPs.
In quantum materials macroscopic behavior is governed in non-trivial ways by quantum phenomena. This is usually achieved by exquisite control over atomic positions in crystalline solids. Here we demonstrate that the use of disordered glassy materials provides unique opportunities to tailor quantum material properties. By borrowing ideas from single molecule spectroscopy, we isolate single delocalized π-electron dye systems in relatively rigid ultrasmall (<10nm diameter) amorphous silica nanoparticles. We demonstrate that chemically tuning the local amorphous silica environment around the dye over a range of compositions enables exquisite control over dye quantum behavior, leading to efficient probes for photodynamic therapy (PDT) and stochastic optical reconstruction microscopy (STORM). Results suggest that efficient fine-tuning of light-induced quantum behavior mediated via effects like spin-orbit coupling can be effectively achieved by systematically varying averaged local environments in glassy amorphous materials as opposed to tailoring well-defined neighboring atomic lattice positions in crystalline solids. Resulting nanoprobes have required features proven to enable clinical translation.
In recent years, high-resolution optical imaging in the far field has provided opportunities for alternative approaches to nanocharacterization traditionally dominated by electron and scanning probe microscopies. Here, we report the optical super-resolution imaging of model block copolymer (BCP) thin film surface nanostructures through stochastic optical reconstruction microscopy (STORM). We compare a set of surface-functionalized fluorescent core–shell silica nanoparticles encapsulating two different organic dyes, Cy3 and Cy5, with the corresponding free dyes in STORM. Using various click-type chemistries, these probes are covalently attached to the surface of specific blocks of BCP thin films, enabling selective block labeling and optical visualization. We demonstrate that the enhanced brightness of these particle probes offers distinct advantages over conventional dye labeling, outperforming one of the best STORM dyes available (Cy5).
Multicolor optical super-resolution microscopy (OSRM) describes an emerging set of techniques for the specific labeling of distinct constituents of multicomponent systems with compatible optical probes, elucidating proximity relationships from far-field imaging of diffraction-limited features with nanometer-scale resolution. While such approaches are well established in the study of biological systems, their implementation in materials science has been considerably slower. In large part, this gradual adoption is due to the lack of appropriate OSRM probes that, e.g., by facile mixing or surface modification, enable orthogonal labeling of specific nanostructures in the condensed state, rather than in aqueous conditions as with biology. Here, OSRM probes in the form of ultrasmall (diameters <10 nm) aluminosilicate nanoparticles encapsulating different fluorescent dyes are tailored to visualize both nanodomains of polystyrene-block-poly[(allyl glycidyl ether)-co-(ethylene oxide)] (PS-b-P(AGE-co-EO)) diblock copolymer thin films. Careful design of nanoprobe surface chemical properties facilitates either selective compatibilization with the nonpolar PS matrix or preferential reactivity with surface allyl groups of the hydrophilic P(AGE-co-EO) minority block. Stochastic optical reconstruction microscopy (STORM) of the resulting polymer–inorganic nanocomposite thin films shows nanodomain features of the two chemically dissimilar blocks consistent with atomic force microscopy results. This work paves the way for multiplexed OSRM analysis of polymer nanocomposite bulk structures.
During breast cancer bone metastasis, tumor cells interact with bone microenvironment components including inorganic minerals. Bone mineralization is a dynamic process and varies spatiotemporally as a function of cancer‐promoting conditions such as age and diet. The functional relationship between skeletal dissemination of tumor cells and bone mineralization, however, is unclear. Standard histological analysis of bone metastasis frequently relies on prior demineralization of bone, while methods that maintain mineral are often harsh and damage fluorophores commonly used to label tumor cells. Here, fluorescent silica nanoparticles (SNPs) are introduced as a robust and versatile labeling strategy to analyze tumor cells within mineralized bone. SNP uptake and labeling efficiency of MDA‐MB‐231 breast cancer cells is characterized with cryo‐scanning electron microscopy and different tissue processing methods. Using a 3D in vitro model of marrow‐containing, mineralized bone as well as an in vivo model of bone metastasis, SNPs are demonstrated to allow visualization of labeled tumor cells in mineralized bone using various imaging modalities including widefield, confocal, and light sheet microscopy. This work suggests that SNPs are valuable tools to analyze tumor cells within mineralized bone using a broad range of bone processing and imaging techniques with the potential to increase the understanding of bone metastasis.
Cartesian polarization analysis transforms a set of surface infrared spectra obtained in different geometries into their Cartesian components using a mathematical transform, providing direct insight into the bonding geometry of adsorbed molecules. This technique was extended to uniaxial substrates and used to analyze solution-deposited, self-assembled benzoate and alkanoate monolayers on rutile (110). This analysis resolved a long-standing controversy regarding the existence of paired molecules in benzoate monolayers, showing that two distinct isomers exist within the monolayer: a tilted tetramer, which is paired, and a twisted monomer, which is not. The two isomers are nearly isoenergetic, as shown by analysis of STM images and complementary DFT simulations. Infrared and XPS spectra as well as STM images of heptanoate and octanoate monolayers showed the formation of complete monolayers (as opposed to sparse layers or multilayers); however, the alkyl chains in the monolayer are disordered and loosely packed with a significant density of conformational defectsa stark contrast to the near-crystalline, all-trans alkyl monolayers typically formed on Au and Si surfaces. The high disorder in the alkanoate monolayers was attributed to geometry, as the density of alkanoate binding sites on rutile (110) is 30% less than the density of alkyl monolayers on Si. The high density of gauche defects in alkanoate monolayers was attributed to the small energy difference between the all-trans and single-gauche-defect conformers in isolated alkyl chains. In contrast, strong intermolecular interactions in tight-packed alkyl monolayers on Au and Si surfaces suppress gauche defect formation.
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