Single-Digit Pathogen and Attomolar Detection with the Naked Eye Using Liposome-Amplified Plasmonic Immunoassay.

We introduce an enzyme-free plasmonic immunoassay with a binary (all-or-none) response. The presence of a single pathogen in the sample results in a chemical cascade reaction leading to a large red to dark-blue colorimetric shift visible to the naked eye. The immediate and amplified response is initiated by a triggered breakdown of cysteine-loaded nanoliposomes and subsequent aggregation of plasmonic gold nanoparticles. Our approach enabled visual detection of a single-digit live pathogen of Salmonella, Listeria, and E. coli O157 in water and food samples. Furthermore, the assay allowed a naked-eye detection of target antibody concentrations as low as 6.7 attomolar (600 molecules in 150 µL); six orders of magnitude lower than conventional enzyme-linked immunosorbent assay (ELISA).

Simultaneous detection of ultratrace lead and copper with gold nanoparticles patterned on carbon nanotube thin film.

Highly sensitive detection of a Pb(2+)-Cu(2+) mixture using gold nanoparticles patterned on single-walled carbon nanotube (AuNP-SWCNT) film is reported. The gold nanoparticles were deposited electrochemically on carbon nanotube film using a cyclic voltammetry technique. The film showed a homogeneous size and density that could be easily controlled by the potential scanning cycle and gold precursor concentration. Square wave stripping voltammetry (SWSV) was applied to the simultaneous detection of Pb(2+) and Cu(2+) under optimized conditions. The AuNP-SWCNT electrode exhibited a high increase in sensitivity with a limit of detection of 0.546 ppb (R(2) = 0.984) and 0.613 ppb (R(2) = 0.991) for Pb(2+) and Cu(2+) ions, respectively, in a mixture of Pb(2+)-Cu(2+) solution (S/N = 3, n = 5), and a good linear response in the range from 3.31 ppb to 22.29 ppb. The electrode exhibited high reproducibility in repetitive measurements with a relative standard deviation as low as 4.2% and 2.6% for Pb(2+) and Cu(2+) ions, respectively. An interference study showed that Sb(3+), As(3+), Zn(2+), Ca(2+), and Na(+) ions did not have a significant effect. This study demonstrated an alternative approach to the rapid and reliable detection of heavy metals of environmental interest.

Enzyme kinetic measurements using a droplet-based microfluidic system with a concentration gradient.

In this paper, we propose a microfluidic device that is capable of generating a concentration gradient followed by parallel droplet formation within channels with a simple T-junction geometry. Linear concentration gradient profiles can be obtained based on fluid diffusion under laminar flow. Optimized conditions for generating a linear concentration gradient and parallel droplet formation were investigated using fluorescent dye. The concentration gradient profile under diffusive mixing was dominated by the flow rate at sample inlets, while parallel droplet formation was affected by the channel geometry at both the inlet and outlet. The microfluidic device was experimentally characterized using optimal layout and operating conditions selected through a design process. Furthermore, in situ enzyme kinetic measurements of the ß-galactosidase-catalyzed hydrolysis of resorufin-ß-d-galactopyranoside were performed to demonstrate the application potential of our simple, time-effective, and low sample volume microfluidic device. We expect that, in addition to enzyme kinetics, drug screening and clinical diagnostic tests can be rapidly and accurately performed using this droplet-based microfluidic system.

Patterning of single-walled carbon nanotube films on flexible, transparent plastic substrates.

We report a simple patterning method for single-walled carbon nanotubes (SWCNTs) films on flexible, transparent poly(ethylene terephthalate) using an O(2)-plasma technique in a capacitively coupled plasma (CCP) system. The homogeneous SWCNT films in a large area were fabricated by the vacuum filtration method. The plasma patterning process of SWCNT films includes conventional photolithography and subsequent O(2)-plasma treatment. During the plasma treatment, SWCNTs underneath the patterned photoresist polymer are protected from etching and damage by O(2)-plasma while the exposed SWCNTs are destroyed. The morphological changes and the effect of plasma treatment on the chemical properties of SWCNT films were investigated by scanning electron microscopy and X-ray photoelectron spectroscopy, respectively. The physical properties of SWCNT films such as transparency and conductivity were systematically characterized under various plasma conditions. In an electrochemiluminescence reaction, the SWCNT films patterned by the CCP system-based O(2)-plasma treatment could be used as flexible and transparent electrodes.

Electrochemical patterning of transparent single-walled carbon nanotube films on plastic substrates.

We report a new patterning method for single-walled carbon nanotubes (SWCNTs) films on flexible, transparent poly(ethylene terephthalate) using electrochemical etching in an aqueous electrolyte solution. Electrochemical etching of the SWCNT films patterned with photoresist polymer was accomplished in a three-electrode system, and the electrochemically patterned SWCNT films were then characterized by scanning electron microscopy (SEM) and Raman spectroscopy. The voltammetry curve showed that SWCNTs underwent drastic oxidation above an applied potential of 1.315 V with the generation of gas bubbles, and the oxidation current became constant above 2.6 V due to the mass transfer limit. SEM images showed that the networks of SWCNTs in the area protected with the photoresist polymer had no damage and vivid connections were obvious, while the connections and shapes of SWCNTs in the area exposed to electrochemical etching were indistinct and slightly damaged. In the Raman spectra of the area protected with the photoresist polymer and the exposed SWCNT area, the intensity ratio of the D-line to the G-line increased from 0.077 to 1.136, which indicated that the ordered carbons of the SWCNT film gradually became amorphous carbons due to electrochemical etching. For optimal patterning, the electrochemical etchings of SWCNT films were performed under various conditions (the applied potential, pH of the electrolyte solution, and electrolyte concentration). An applied potential of 3.0 V in 0.1 M NaCl electrolyte solution (pH 7.0) was optimal for homogeneous electrochemical patterning of SWCNT films. In an electrochemiluminescence reaction, the SWCNT films patterned by this technique could be used successfully as flexible and transparent electrodes.

Metal-organic polyhedron based on a Cu(II) paddle-wheel secondary building unit at the truncated octahedron corners.

A metal-organic polyhedron of truncated octahedral geometry augmented with a C(4)-symmetric square-planar Cu(II) paddle-wheel node as a secondary building unit can be prepared using a C(3)-symmetric ligand that occupies the face of the octahedral cage, where the three phenyl groups containing a m-carboxylate group in the ligand provide the necessary curvature to form the finite octahedral cage.

Gold Nanoplate-Enhanced Chemiluminescence and Macromolecular Shielding for Rapid Microbial Diagnostics.

With the global rise of antimicrobial resistance, rapid screening and identification of low concentrations of microorganisms in less than 1 h becomes an urgent technological need for evidence-based antibiotic therapy. Although many commercially available techniques are labeled for rapid microbial detection, they often require 24-48 h of cell enrichment to reach detectable levels. Here, it is shown that the widely used reducing agent tris(2-carboxyethyl)phosphine (TCEP) can also act as a powerful oxidant on gold nanoplates and subsequently lead to a strong catalysis of luminol chemiluminescence. The catalytic reaction results in up to 100-fold signal enhancement and unprecedented stable luminescence for up to 10 min. However, when TCEP is exposed to microorganisms, it is oxidized by the microbial surface proteins and loses its catalytic properties, leading to a decrease in chemiluminescence. The competitive interaction of TCEP with Au nanoplates and microorganisms is used to introduce a homogenous rapid detection method that allows microbial screening in less than 10 min with a limit of detection down to 100 cfu mL-1 . Furthermore, the concept of microbial macromolecular shielding using antibody-conjugated polymers is introduced. The combination of TCEP redox activity and macromolecular shielding enables specific microbial identification within 1 h, without preconcentration, cell enrichment, or heavy equipment other than a hand-held luminometer. The technique is demonstrated by specific detection of methicillin-resistant Staphylococcus aureus in environmental and urine samples containing a mixture of microorganisms.

Paper-based chemical and biological sensors: Engineering aspects.

Remarkable efforts have been dedicated to paper-based chemosensors and biosensors over the last few years, mainly driven by the promise of reaching the best trade-off between performance, affordability and simplicity. Because of the low-cost and rapid prototyping of these sensors, recent research has been focused on providing affordable diagnostic devices to the developing world. The recent progress in sensitivity, multi-functionality and integration of microfluidic paper-based analytical devices (µPADs), increasingly suggests that this technology is not only attractive in resource-limited environments but it also represents a serious challenger to silicon, glass and polymer-based biosensors. This review discusses the design, chemistry and engineering aspects of these developments, with a focus on the past few years.

Microgel-encapsulated methylene blue for the treatment of breast cancer cells by photodynamic therapy.

PURPOSE: Photodynamic therapy (PDT) is gaining increasing recognition for breast cancer treatment because it offers local selectivity and reduced toxic side effects compared to radiotherapy and chemotherapy. In PDT, photosensitizer drugs are loaded in different nanomaterials and used in combination with light exposure. However, the most representative issue with PDT is the difficulty of nanomaterials to encapsulate anticancer drugs at high doses, which results in low efficacy of the PDT treatment. Here, we proposed the development of the poly(N-isopropylacrylamide) (PNIPAM) microgel for the encapsulation of methylene blue, an anticancer drug, for its use as breast cancer treatment in MCF-7 cell line. METHODS: We developed biocompatible microgels based on nonfunctionalized PNIPAM and its corresponding anionically functionalized PNIPAM and polyacrylic acid (PNIPAM-co-PAA) microgel. Methylene blue was used as the photosensitizer drug because of its ability to generate toxic reactive oxygen species upon exposure to light at 664 nm. Core PNIPAM and core/shell PNIPAM-co-PAA microgels were synthesized and characterized using ultraviolet-visible spectroscopy and dynamic light scattering. The effect of methylene blue was evaluated using the MCF-7 cell line. RESULTS: Loading of methylene blue in core PNIPAM microgel was higher than that in the core/shell PNIPAM-co-PAA microgel, indicating that electrostatic interactions did not play an important role in loading a cationic drug. This behavior is probably due to the skin layer inhibiting the high uptake of drugs in the PNIPAM-co-PAA microgel. Core PNIPAM microgel effectively retained the cationic drug (i.e., methylene blue) for several hours compared to core/shell PNIPAM-co-PAA and enhanced its photodynamic efficacy in vitro more than that of free methylene blue. CONCLUSION: Our results showed that the employment of core PNIPAM and core/shell PNIPAM-co-PAA microgels enhanced the encapsulation of methylene blue. Core PNIPAM microgel released the drug more slowly than did core/shell PNIPAM-co-PAA, and it effectively inhibited the growth of MCF-7 cells.

Electrochemical determination of cadmium and lead on pristine single-walled carbon nanotube electrodes.

A flexible, transparent, single-walled carbon nanotube (SWCNT) film electrode was prepared by vacuum filtering methods, followed by photolithographic patterning of a photoresist polymer on the SWCNT surface. The morphology of the SWCNT film electrode surface was characterized using a field-emission scanning electron microscope coupled to an energy-dispersive X-ray spectrophotometer. The electrodes were successfully used as a mercury-free electrochemical sensor for individual and simultaneous detection of cadmium (Cd(2+)) and lead (Pb(2+)) in 0.02 M HCl by square-wave stripping voltammetry. Some important operational parameters, including deposition time, deposition potential, square-wave amplitude, and square wave-frequency were optimized for the detection of Cd(2+) and Pb(2+). The newly developed sensor showed good linear behavior in the examined concentration. For individual Cd(2+) and Pb(2+) ion detection, the linear range was found from 0.033 to 0.228 ppm with detection limits of 0.7 ppb (R(2) = 0.985) for Cd(2+) and 0.8 ppb (R(2) = 0.999) for Pb(2+). For simultaneous detection, the linear range was found from 0.033 to 0.280 ppm with a limit of detection of 2.2 ppb (R(2) = 0.976) and 0.6 ppb (R(2) = 0.996) for Cd(2+) and Pb(2+), respectively. SWCNT film electrodes offered favorable reproducibility of ± 5.4% and 4.3% for Cd(2+) and Pb(2+), respectively. The experiments demonstrated the applicability of carbon nanotubes, specifically in the preparation of SWCNT films. The results suggest that the proposed flexible SWCNT film electrodes can be applied as simple, efficient, cost-effective, and/or disposable electrodes for simultaneous detection of heavy metal ions.

Control of ZnO morphologies on carbon nanotube electrodes and electrocatalytic characteristics toward hydrazine.

We controlled the morphologies of zinc oxide (ZnO) nanostructures on single-walled carbon nanotube electrodes by an electrochemical deposition method and investigated the dependence of the electrocatalytic characteristics toward hydrazine on the different morphologies. ZnO nanorods provided high electrocatalytic activity with unique electrochemical behaviours, associated with the H(+) ion generated by the electro-oxidation of hydrazine.

Electrochemical characterization of a single-walled carbon nanotube electrode for detection of glucose.

We developed glucose biosensing electrodes using single-walled carbon nanotube (SWCNT) films on flexible, transparent poly(ethylene terephthalate). The homogeneous SWCNT films were fabricated by a vacuum filtration method, and the averaged resistivity and transparency of the fabricated flexible SWCNT films were 400 Omega sq(-1) and 80%, respectively. The glucose sensing electrodes were constructed by encapsulating glucose oxidase (GOx) by Nafion binder into the SWCNT film, and the variation in current response as a function of enzyme loading amount, Nafion thickness were investigated. 30 mg mL(-1) GOx and 2% Nafion was optimal for the detection of glucose. When ferrocene monocarboxylic acid (FMCA) was introduced as diffusional electron mediator, the current responses toward glucose of the Nafion/GOx/SWCNT electrodes in glucose solution containing FMCA were dramatically improved, and the developed sensor was independent of oxygen. In the application of GOx immobilized SWCNT films for glucose detection, a linear electrical response was observed for concentrations ranging from 0.25 to 3.0 mM, and the detection limit and the sensitivity were assessed to be 97 microM and 9.32 microA mM(-1) cm(-2), respectively. Moreover, according to the Lineweaver-Burk plot, the apparent Michaelis-Menten constant was calculated to be 23.8 mM, and the current responses did not interfere with coexisting electroactive species, indicating that Nafion is an effective permselective polymer barrier.

Electrochemical patterning of gold nanoparticles on transparent single-walled carbon nanotube films.

We report a simple, low cost, electrochemical deposition method to pattern gold nanoparticles on flexible, transparent, single-walled carbon nanotube (SWCNT) films, and demonstrate the application of the gold-patterned SWCNT films as surface-enhanced Raman spectroscopy substrates and biosensing electrodes for non-enzymatic glucose detection.

Preparation of monodisperse and size-controlled poly(ethylene glycol) hydrogel nanoparticles using liposome templates.

Liposomes were used as templates to prepare size-controlled and monodisperse poly(ethylene glycol) (PEG) hydrogel nanoparticles. The procedure for the preparation of PEG nanoparticles using liposomes consists of encapsulation of photopolymerizable PEG hydrogel solution into the cavity of the liposomes, extrusion through a membrane with a specific pore size, and photopolymerization of the contents inside the liposomes by UV irradiation. The size distributions of the prepared particles were 1.32+/-0.16 microm (12%), 450+/-62 nm (14%), and 94+/-12 nm (13%) after extrusion through membrane filters with pore sizes of 1 microm, 400 nm, and 100 nm, respectively. With this approach, it is also possible to modify the surface of the hydrogel nanoparticles with various functional groups in a one-step procedure. To functionalize the surface of a PEG nanoparticle, methoxy poly(ethylene glycol)-aldehyde was added as copolymer to the hydrogel-forming components and aldehyde-functionalized PEG nanoparticles could be obtained easily by UV-induced photopolymerization, following conjugation with poly-L-lysine-FITC through amine-aldehyde coupling. The prepared PEG particles showed strong fluorescence from FITC on the edge of the particles using confocal microscopy. The immobilization of biomaterials such as enzymes in hydrogel particles could be performed with loading beta-galactosidases during the hydration step for liposome preparation without additional procedures. The resorufin produced by applying resorufin beta-D-galactopyranoside as the substrate showed the fluorescence under the confocal microscopy.

Gold nanoparticle aggregation-based highly sensitive DNA detection using atomic force microscopy.

The potential ability of atomic force microscopy (AFM) as a quantitative bioanalysis tool is demonstrated by using gold nanoparticles as a size enhancer in a DNA hybridization reaction. Two sets of probe DNA were functionalized on gold nanoparticles and sandwich hybridization occurred between two probe DNAs and target DNA, resulting in aggregation of the nanoparticles. At high concentrations of target DNA in the range from 100 nM to 10 microM, the aggregation of gold nanoparticles was determined by monitoring the color change with UV-vis spectroscopy. The absorption spectra broadened after the exposure of DNA-gold nanoparticles to target DNA and a new absorption band at wavelengths >600 nm was observed. However, no differences were observed in the absorption spectra of the gold nanoparticles at low concentrations of target DNA (10 pM to 10 nM) due to insufficient aggregation. AFM was used as a biosensing tool over this range of target DNA concentrations in order to monitor the aggregation of gold nanoparticles and to quantify the concentration of target DNA. Based on the AFM images, we successfully evaluated particle number and size at low concentrations of target DNA. The calibration curve obtained when mean particle aggregate diameter was plotted against concentration of target DNA showed good linearity over the range 10 pM to 10 nM, the working range for quantitative target DNA analysis. This AFM-based DNA detection technique was three orders of magnitude more sensitive than a DNA detection method based on UV-vis spectroscopy.