Radiation-Tolerant Thin-Film Electrodes for pH Monitoring in Sterile Media.

Thin-film pH electrodes on thermoplastic substrates can be subjected to γ-radiation (up to 45 kGy) without loss of stability or sensing performance, with important ramifications for monitoring analytes in sterile environments. pH-sensing membranes composed of polyvinyl chloride (PVC), trioctyl trimellitate (TOTM), and a standard hydrogen ionophore were cast onto screen-printed carbon electrodes with exfoliated graphene as a solid contact. Irradiated thin-film electrodes were conditioned in phosphate buffers and monitored for up to 3 months for changes in voltage readout and pH sensitivity, relative to untreated controls. The sensitivities of both irradiated and control electrodes were consistently Nernstian over a 100 day window, with both types exhibiting logarithmic voltage decays but in opposite directions. The γ-irradiated electrodes had excellent long-term stability with quasi-linear voltage drifts of +0.28 mV (∼0.005 pH) per day. Voltage readouts from sterilized thin-film electrodes in cell culture media could be converted by single-point calibration into pH values that fell within 0.07 units relative to a commercial pH meter (calibrated daily).

Label-Free Detection and Discrimination of Bacterial Pathogens Based on Hemin Recognition.

Hemin linked to hexa(ethylene glycol)bishydrazide was patterned by inkjet printing into periodic microarrays, and evaluated for their ability to capture bacterial pathogens expressing various hemin receptors. Bacterial adhesion was imaged under darkfield conditions with Fourier analysis, supporting a label-free method of pathogen detection. Hemin microarrays were screened against a panel of 16 bacteria and found capable of capturing multiple species, some with limits of detection as low as 10(3) cfu/mL. Several Gram-positive strains including Staphylococcus aureus and Bacillus anthracis also exhibited rapid adhesion, enabling pattern recognition within minutes of exposure. This can be attributed to differences in hemin acquisition systems: aggressively adherent bacteria express cell-surface hemin receptors (CSHRs) that enable direct hemin binding and uptake, whereas other types of bacteria including most Gram-negative strains rely on the secretion and recapture of soluble proteins (hemophores) for hemin acquisition, with consequently longer times for ligand binding and detection.

Cys34-PEGylated Human Serum Albumin for Drug Binding and Delivery.

Polyethylene glycol (PEG) derivatives were conjugated onto the Cys-34 residue of human serum albumin (HSA) to determine their effects on the solubilization, permeation, and cytotoxic activity of hydrophobic drugs such as paclitaxel (PTX). PEG(C34)HSA conjugates were prepared on a multigram scale by treating native HSA (n-HSA) with 5- or 20-kDa mPEG-maleimide, resulting in up to 77% conversion of the mono-PEGylated adduct. Nanoparticle tracking analysis of PEG(C34)HSA formulations in phosphate buffer revealed an increase in the number of nanosized aggregates relative to n-HSA, both in the absence and presence of PTX. Cell viability studies conducted with MCF-7 breast cancer cells indicated that PTX cytotoxicity was enhanced by PEG(C34)HSA when mixed at 10:1 mol ratios, up to a 2-fold increase in potency relative to n-HSA. The PEG(C34)HSA conjugates were also evaluated as PTX carriers across monolayers of HUVEC and hCMEC/D3 cells, and found to have permeation profiles nearly identical to those of n-HSA.

Nanometric resolution in the hydrodynamic size analysis of ligand-stabilized gold nanorods.

The stability and hydrodynamic size of ligand-coated gold nanorods (GNRs; aspect ratio 3.6) have been characterized by nanoparticle tracking analysis (NTA)-a single-particle counting method that can measure size distributions with low nanometer resolution. Stable aqueous suspensions of citrate-stabilized GNRs (cit-GNRs) are amenable to surface functionalization without loss of dispersion control. Cit-GNRs can be treated with chemisorptive ligands (thiols and dithiocarbamates), nonionic surfactants (Tween 20), and proteins (human serum albumin), all of which produce stable suspensions at low surfactant concentrations. The precision of NTA (relative standard deviation 10-12%, standard error <2%) is sufficient to allow differences in the hydrodynamic size of coated GNRs to be interpreted in terms of surfactant structure and conformation.

Antidelaminating, Thermally Stable, and Cost-Effective Flexible Kapton Platforms for Nitrate Sensors, Mercury Aptasensors, Protein Sensors, and p-Type Organic Thin-Film Transistors.

The inkjet printing of metal electrodes on polymer films is a desirable manufacturing process due to its simplicity but is limited by the lack of thermal stability and serious delaminating flaws in various aqueous and organic solutions. Kapton, adopted worldwide due to its superior thermal durability, allows the high-temperature sintering of nanoparticle-based metal inks. By carefully selecting inks (Ag and Au) and Kapton substrates (Kapton HN films with a thickness of 135 µm and a thermal resistance of up to 400 °C) with optimal printing parameters and simplified post-treatments (sintering), outstanding film integrity, thermal stability, and antidelaminating features were obtained in both aqueous and organic solutions without any pretreatment strategy (surface modification). These films were applied in four novel devices: a solid-state ion-selective (IS) nitrate (NO3-) sensor, a single-stranded DNA (ssDNA)-based mercury (Hg2+) aptasensor, a low-cost protein printed circuit board (PCB) sensor, and a long-lasting organic thin-film transistor (OTFT). The IS NO3- sensor displayed a linear sensitivity range between 10-4.5 and 10-1 M (r2 = 0.9912), with a limit of detection of 2 ppm for NO3-. The Hg2+ sensor exhibited a linear correlation (r2 = 0.8806) between the change in the transfer resistance (RCT) and the increasing concentration of Hg2+. The protein PCB sensor provided a label-free method for protein detection. Finally, the OTFT demonstrated stable performance, with mobility values in the linear (µlin) and saturation (µsat) regimes of 0.0083 ± 0.0026 and 0.0237 ± 0.0079 cm2 V-1 S-1, respectively, and a threshold voltage (Vth) of -6.75 ± 3.89 V.

Polymer-iron oxide composite nanoparticles for EPR-independent drug delivery.

Nanoparticle (NP)-based approaches to cancer drug delivery are challenged by the heterogeneity of the enhanced permeability and retention (EPR) effect in tumors and the premature attrition of payload from drug carriers during circulation. Here we show that such challenges can be overcome by a magnetophoretic approach to accelerate NP delivery to tumors. Payload-bearing poly(lactic-co-glycolic acid) NPs were converted into polymer-iron-oxide nanocomposites (PINCs) by attaching colloidal Fe3O4 onto the surface, via a simple surface modification method using dopamine polymerization. PINCs formed stable dispersions in serum-supplemented medium and responded quickly to magnetic field gradients above 1 kG/cm. Under the field gradients, PINCs were rapidly transported across physical barriers and into cells and captured under flow conditions similar to those encountered in postcapillary venules, increasing the local concentration by nearly three orders of magnitude. In vivo magnetophoretic delivery enabled PINCs to accumulate in poorly vascularized subcutaneous SKOV3 xenografts that did not support the EPR effect. In vivo magnetic resonance imaging, ex vivo fluorescence imaging, and tissue histology all confirmed that the uptake of PINCs was higher in tumors exposed to magnetic field gradients, relative to negative controls.

Uniform gold nanorod arrays from polyethylenimine-coated alumina templates.

Monolithic Au nanorod arrays can be grown by electrodeposition in Au-backed nanoporous alumina templates using polyethylenimine (PEI) as an adhesion layer, with excellent height control between 300 nm and 1.4 microm. The local height distribution can be extremely narrow with relative standard deviations well below 2%. The uniform growth rate appears to be determined by the adsorbed PEI matrix, which controls the growth kinetics of the grains comprising the nanorods. The nanorods can be retained as free-standing 2D arrays after careful removal of the AAO template. Reflectance spectroscopy reveals a collective plasmon mode with a maximum near 1.2 microm, in accord with recent calculations for 2D arrays of closely spaced cylindrical nanoparticles.

Nanoprobe implantation into mammalian cells by cationic transfection.

Submicron-sized Au particles and Au/SiO(2) nanocomposites (superparticles) as large as 670 nm have been introduced into tsA201 cells with minimal cell trauma by cationic transfection systems. Successful implantations can be characterized by the expression of co-transfected DNA.

Preparation of Super-Stable Gold Nanorods via Encapsulation into Block Copolymer Micelles.

Gold nanorods (GNRs) have the potential to be used as imaging and hyperthermia agents for cancer theranostics. Clinical applications of as-synthesized GNRs (i.e., cetyl trimethylammonium bromide (CTAB)-coated GNRs) are currently limited by their cytotoxicity and insufficient colloidal stability. With an aim to address these problems, we developed a self-assembly processing technique for encapsulating GNRs in poly(ethylene oxide)-poly(n-butyl acrylate) (PEO-PnBA) block copolymer (BCP) micelles. This technique uses simple steps of solvent exchange processes, based on the known principles of block copolymer self-assembly. The resultant BCP-encapsulated GNRs were found to be stable against aggregation under physiological salt conditions for indefinite periods of time, which has rarely previously been achieved by other means of encapsulation.

Gold nanorods: multifunctional agents for cancer imaging and therapy.

Gold nanorods (GNRs) are strongly absorbing at near-infrared (NIR) frequencies and can be employed as multifunctional agents for biological imaging and theragnostics. GNRs can support nonlinear optical microscopies based on two-photon-excited luminescence and can enhance the contrast of biomedical imaging modalities such as optical coherence tomography and photoacoustic tomography. GNRs are also efficient at mediating the conversion of NIR light energy into heat and can generate localized photothermal effects. However, future clinical applications will require the rigorous removal of CTAB, a micellar surfactant used in GNR synthesis, and reliable methods of surface functionalization for cell-selective targeting and for minimizing nonspecific uptake into cells. This can be accomplished by using polystyrenesulfonate (PSS) as a sorbent for removing CTAB, and in situ dithiocarbamate formation for introducing chemisorptive ligands onto GNR surfaces.

Detoxification of gold nanorods by treatment with polystyrenesulfonate.

We address an outstanding issue associated with the biocompatibility of gold nanorods (GNRs), a promising agent for biomedical imaging and theragnostics. GNRs are typically prepared in the presence of cetyltrimethylammonium bromide (CTAB), a cationic surfactant whose rigorous removal is necessary due to its cytotoxicity and membrane-compromising properties. CTAB-stabilized GNRs can be partially purified by treatment with polystyrenesulfonate (PSS), an anionic polyelectrolyte often used as a surrogate peptizing agent, followed by chloroform extraction and ultrafiltration with minimal loss of dispersion stability. However, in vitro cytotoxicity assays of PSS-coated GNRs revealed IC(50) values in the low to submicromolar range, with subsequent studies indicating the source of toxicity to be associated with a persistent PSS-CTAB complex. Further exchange of CTAB-laden PSS with fresh polyelectrolyte greatly improves biocompatibility, to the extent that 85 microg/mL of "CTAB-free" GNRs (the highest level evaluated) has comparable toxicity to a standard phosphate buffer solution. Ironically, PSS is not effective by itself at stabilizing GNRs in CTAB-depleted suspensions: while useful as a detergent for GNR detoxification, it should be replaced by more robust coatings for long-term stability under physiological conditions.

Controlling the cellular uptake of gold nanorods.

Gold nanorods coated with cetyltrimethylammonium bromide (CTAB), a cationic micellar surfactant used in nanorod synthesis, were rapidly and irreversibly internalized by KB cells via a nonspecific uptake mechanism. Internalized nanorods near the cell surface were monitored by two-photon luminescence (TPL) microscopy and observed to migrate toward the nucleus with a quadratic rate of diffusion. The internalized nanorods were not excreted but formed permanent aggregates within the cells, which remained healthy and grew to confluence over a 5-day period. Nonspecific nanorod uptake could be greatly reduced by displacing the CTAB surfactant layer with chemisorptive surfactants, particularly by the conjugation of poly(ethylene glycol) chains onto nanorods using in situ dithiocarbamate formation.