A Comparison of Iron Oxide Particles and Silica Particles for Tracking Organ Recellularization.

Reseeding of decellularized organ scaffolds with a patient's own cells has promise for eliminating graft versus host disease. This study investigated whether ultrasound imaging or magnetic resonance imaging (MRI) can track the reseeding of murine liver scaffolds with silica-labeled or iron-labeled liver hepatocytes. Mesoporous silica particles were created using the Stöber method, loaded with Alexa Flour 647 fluorophore, and conjugated with protamine sulfate, glutamine, and glycine. Fluorescent iron oxide particles were obtained from a commercial source. Liver cells from donor mice were loaded with the silica particles or iron oxide particles. Donor livers were decellularized and reperfused with silica-labeled or iron-labeled cells. The reseeded livers were longitudinally analyzed with ultrasound imaging and MRI. Liver biopsies were imaged with confocal microscopy and scanning electron microscopy. Ultrasound imaging had a detection limit of 0.28 mg/mL, while MRI had a lower detection limit of 0.08 mg/mL based on particle weight. The silica-loaded cells proliferated at a slower rate compared to iron-loaded cells. Ultrasound imaging, MRI, and confocal microscopy underestimated cell numbers relative to scanning electron microscopy. Ultrasound imaging had the greatest underestimation due to coarse resolution compared to the other imaging modalities. Despite this underestimation, both ultrasound imaging and MRI successfully tracked the longitudinal recellularization of liver scaffolds.

Hybrid phospholipid bilayer coatings for separations of cationic proteins in capillary zone electrophoresis.

Protein separations in CZE suffer from nonspecific adsorption of analytes to the capillary surface. Semipermanent phospholipid bilayers have been used to minimize adsorption, but must be regenerated regularly to ensure reproducibility. We investigated the formation, characterization, and use of hybrid phospholipid bilayers (HPBs) as more stable biosurfactant capillary coatings for CZE protein separations. HPBs are formed by covalently modifying a support with a hydrophobic monolayer onto which a self-assembled lipid monolayer is deposited. Monolayers prepared in capillaries using 3-cyanopropyldimethylchlorosilane (CPDCS) or n-octyldimethylchlorosilane (ODCS) yielded hydrophobic surfaces with lowered surface free energies of 6.0 ± 0.3 or 0.2 ± 0.1 mJ m(-2) , respectively, compared to 17 ± 1 mJ m(-2) for bare silica capillaries. HPBs were formed by subsequently fusing vesicles comprised of 1,2-dilauroyl-sn-glycero-3-phosphocholine or 1,2-dioleoyl-sn-glycero-3-phosphocholine to CPDCS- or ODCS-modified capillaries. The resultant HPB coatings shielded the capillary surface and yielded reduced electroosmotic mobility (1.3-1.9 × 10(-4) cm(2) V(-1) s(-1) ) compared to CPDCS- and ODCS-modified or bare capillaries (3.6 ± 0.2 × 10(-4) cm(2) V(-1) s(-1) , 4.8 ± 0.4 × 10(-4) cm(2) V(-1) s(-1) , and 6.0 ± 0.2 × 10(-4) cm(2) V(-1) s(-1) , respectively), with increased stability compared to phospholipid bilayer coatings. HPB-coated capillaries yielded reproducible protein migration times (RSD ≤ 3.6%, n ≥ 6) with separation efficiencies as high as 200 000 plates/m.

Hybrid Nanoparticle Platform for Nanoscale Scintillation Proximity Assay.

ß-particle emitting radionuclides, such as 3H, 14C, 32P, 33P, and 35S, are important molecular labels due to their small size and the prevalence of these atoms in biomolecules but are challenging to selectively detect and quantify within aqueous biological samples and systems. Here, we present a core-shell nanoparticle-based scintillation proximity assay platform (nanoSPA) for the separation-free, selective detection of radiolabeled analytes. nanoSPA is prepared by incorporating scintillant fluorophores into polystyrene core particles and encapsulating the scintillant-doped cores within functionalized silica shells. The functionalized surface enables covalent attachment of specific binding moieties such as small molecules, proteins, or DNA that can be used for analyte-specific detection. nanoSPA was demonstrated for detection of 3H-labeled analytes, the most difficult biologically relevant ß-emitter to measure due to the low energy ß-particle emission, using three model assays that represent covalent and non-covalent binding systems that necessitate selectivity over competing 3H-labeled species. In each model, nmol quantities of target were detected directly in aqueous solution without separation from unbound 3H-labeled analyte. The nanoSPA platform facilitated measurement of 3H-labeled analytes directly in bulk aqueous samples without surfactants or other agents used to aid particle dispersal. Selectivity for bound 3H-analytes over unbound 3H analytes was enhanced up to 30-fold when the labeled species was covalently bound to nanoSPA, and 4- and 8-fold for two non-covalent binding assays using nanoSPA. The small size and enhanced selectivity of nanoSPA should enable new applications compared to the commonly used microSPA platform, including the potential for separation-free, analyte-specific cellular or intracellular detection.

Polystyrene-Core, Silica-Shell Scintillant Nanoparticles for Low-Energy Radionuclide Quantification in Aqueous Media.

ß-particle emitting radionuclides are useful molecular labels due to their abundance in biomolecules. Detection of ß-emission from 3H, 35S, and 33P, important biological isotopes, is challenging due to the low energies (Emax ≤ 300 keV) and short penetration depths (≤0.6 mm) in aqueous media. The activity of biologically relevant ß-emitters is usually measured in liquid scintillation cocktail (LSC), a mixture of energy-absorbing organic solvents, surfactants, and scintillant fluorophores, which places significant limitations on the ability to acquire time-resolved measurements directly in aqueous biological systems. As an alternative to LSC, we developed polystyrene-core, silica-shell nanoparticle scintillators (referred to as nanoSCINT) for quantification of low-energy ß-particle emitting radionuclides directly in aqueous solutions. The polystyrene acts as an absorber for energy from emitted ß-particles and can be loaded with a range of hydrophobic scintillant fluorophores, leading to photon emission at visible wavelengths. The silica shell serves as a hydrophilic shield for the polystyrene core, enabling dispersion in aqueous media and providing better compatibility with water-soluble analytes. While polymer and inorganic scintillating microparticles are commercially available, their large size and/or high density complicates effective dispersion throughout the sample volume. In this work, nanoSCINT nanoparticles were prepared and characterized. nanoSCINT responds to 3H, 35S, and 33P directly in aqueous solutions, does not exhibit a change in scintillation response between pH 3.0 and 9.5 or with 100 mM NaCl, and can be recovered and reused for activity measurements in bulk aqueous samples, demonstrating the potential for reduced production of LSC waste and reduced total waste volume during radionuclide quantification. The limits of detection for 1 mg/mL nanoSCINT are 130 nCi/mL for 3H, 8 nCi/mL for 35S, and <1 nCi/mL for 33P.

Fate of fluorescent core-shell silica nanoparticles during simulated secondary wastewater treatment.

Increasing use of silica nanoparticles (SiO2 NPs) in consumer products and industrial processes leads to SiO2 NP discharge into wastewater. Thus, there is a need to understand the fate of SiO2 NPs during wastewater treatment. However, the detection of SiO2 NPs in environmental systems is hindered by the elevated background levels of natural silicon. In this work, laboratory-synthesized fluorescent core-shell SiO2 NPs were used to study the fate of these NPs during secondary wastewater treatment. Fluorescent measurements provided an easy and fast method for SiO2 NP tracking. A laboratory-scale activated sludge system consisting of an aeration tank and a settler was fed with synthetic wastewater containing ca. 7.5 mg L(-1) of fluorescent SiO2 NPs for 30 days. SiO2 NPs were effectively removed from the wastewater (>96%) during the first 6 days, however the concentration of SiO2 NPs in the effluent gradually increased afterwards and the NP discharge was as high as 65% of the input after 30 days of NP dosing. The poor removal of the SiO2 NPs was related to the high colloidal stability of the NPs in the wastewater and their limited propensity to biosorption. Although some degree of NP adsorption on the biomass was observed using fluorescence microscopy, the affinity of SiO2 NPs for the activated sludge was not enough for a sustained and effective removal of the SiO2 NPs from the wastewater.