Enrichment of adenosine using thermally responsive chromatographic materials under friendly pH conditions

A thermally responsive boronate affinity chromatographic material, which showed thermal sensitivity, had been successfully applied for the enrichment and separation of cis‐diol‐containing compounds, and the capture and release process could be facilitated by adjusting the temperature. However, in this system, the pH of the mobile phase must be higher than 9.8, and alkaline media can lead to the degradation of labile compounds; the use of silica beads also limits its use. In this study, thermally responsive boronate affinity chromatographic material, namely poly(N‐isopropylacrylamide‐co‐N‐acryloyl‐3‐aminophenylboronic acid) grafted silica, was successfully prepared by atom transfer radical polymerization. Its structure was confirmed by IR spectroscopy and the graft ratio was 20.8%, determined by thermogravimetric analysis. Furthermore, the capture/release of adenosine, a cis‐diol, was performed from pH 5.0–9.0 and 10–50°C. The elution of adenosine was remarkably retarded at decreased temperatures and adenosine could be captured completely at 10°C at pH values of 5.0–9.0. The enrichment of adenosine could be achieved by simply changing the temperature from 10 to 50°C. Therefore, this material not only improved the stability of the silica, but was also suitable for the capture of oxidation‐sensitive biological analytes. Moreover, it could be used for the enrichment of cis‐diol‐containing compounds in LC with MS.     

Fabrication of large-area ferromagnetic arrays using etched nanosphere lithography

Nanosphere lithography (NSL) is a simple, cost-effective, and powerful technique capable of producing large-area arrays of ferromagnetic nanostructures with dimensions below 100 nm. These properties make NSL an attractive process for the fabrication of arrays of magnetic elements with applications in magnetic data storage. The main disadvantage with conventional NSL is that the monolayer of spheres always contains imperfections that are transferred to the resulting nanostructures. This can significantly affect the structural and magnetic properties of the fabricated array. In this paper we present a novel adaptation of NSL that reduces the effect of such defects on the resulting nanostructures. The technique also offers excellent control over the diameter, aspect ratio, and pitch of the fabricated elements. These properties are demonstrated through the fabrication of arrays of Ni elements of 210 nm diameter and arrays of Co elements with diameters between 200 and 320 nm.     

Synthesis and aggregation behavior of thermally responsive star polymers

To mimic the three-dimensional (3-D) globular architecture resulting from the precise positioning of hydrophobic/hydrophilic domains (blocks) of naturally occurring proteins, water-soluble linear and star homopolymers of N,N'-dimethylacrylamide (DMA) were synthesized with prescribed molecular weights via reversible addition-fragmentation chain transfer (RAFT) polymerization and subsequently used as macro chain transfer agents for block copolymerization with N-isopropylacrylamide (NIPAM). For the star block copolymers, the interior block consisted of NIPAM while the exterior block was DMA. Since polyNIPAM thermally switches from hydrophilic to hydrophobic, the 3-D solution conformations of the polymers were studied as a function of temperature using differential scanning calorimetry (DSC), static light scattering (SLS), and dynamic light scattering (DLS). The polymers were observed to form monodisperse aggregates in an aqueous pH 4 buffer solution when heated above the lower critical solution temperature (LCST) of polyNIPAM. The temperature at which the polymers aggregated and the size of the aggregates were dependent on the NIPAM block length and the core architecture. A simple model based on an optimal area per headgroup was used to analyze our experimental findings and was useful for predicting the final size and molecular weight of the aggregates formed.     

Fabrication of highly ordered TiO2 nanotube arrays using an organic electrolyte

Anodization of titanium in a fluorinated dimethyl sulfoxide (DMSO) and ethanol mixture electrolyte is investigated. The prepared anodic film has a highly ordered nanotube-array surface architecture. Using a 20 V anodization potential (vs Pt) nanotube arrays having an inner diameter of 60 nm and 40 nm wall thickness are formed. The overall length of the nanotube arrays is controlled by the duration of the anodization, with nanotubes appearing only after approximately 48 h; a 72 h anodization results in a nanotube array approximately 2.3 mum in length. The photoelectrochemical response of the nanotube-array photoelectrodes is studied using a 1 M KOH solution under both UV and visible (AM 1.5) illumination. Enhanced photocurrent density is observed for samples obtained in the organic electrolyte, with an UV photoconversion efficiency of 10.7%.     

Fabrication of hybrid nanostructured arrays using a PDMS/PDMS replication process

In the study, a novel and low cost nanofabrication process is proposed for producing hybrid polydimethylsiloxane (PDMS) nanostructured arrays. The proposed process involves monolayer self-assembly of polystyrene (PS) spheres, PDMS nanoreplication, thin film coating, and PDMS to PDMS (PDMS/PDMS) replication. A self-assembled monolayer of PS spheres is used as the first template. Second, a PDMS template is achieved by replica moulding. Third, the PDMS template is coated with a platinum or gold layer. Finally, a PDMS nanostructured array is developed by casting PDMS slurry on top of the coated PDMS. The cured PDMS is peeled off and used as a replica surface. In this study, the influences of the coating on the PDMS topography, contact angle of the PDMS slurry and the peeling off ability are discussed in detail. From experimental evaluation, a thickness of at least 20 nm gold layer or 40 nm platinum layer on the surface of the PDMS template improves the contact angle and eases peeling off. The coated PDMS surface is successfully used as a template to achieve the replica with a uniform array via PDMS/PDMS replication process. Both the PDMS template and the replica are free of defects and also undistorted after demoulding with a highly ordered hexagonal arrangement. In addition, the geometry of the nanostructured PDMS can be controlled by changing the thickness of the deposited layer. The simplicity and the controllability of the process show great promise as a robust nanoreplication method for functional applications.     

Fabrication of immunosensor microwell arrays from gold compact discs for detection of cancer biomarker proteins

A simple method is reported to fabricate gold arrays featuring microwells surrounding 8-electrodes from gold compact discs (CDs) for less than $0.2 per chip. Integration of these disposable gold CD array chips with microfluidics provided inexpensive immunoarrays that were used to measure a cancer biomarker protein quickly at high sensitivity. The gold CD sensor arrays were fabricated using thermal transfer of laserjet toner from a computer-printed pattern followed by selective chemical etching. Sensor elements had an electrochemically addressable surface area of 0.42 mm(2) with RSD <2%. For a proof-of-concept application, the arrays were integrated into a simple microfluidic device for electrochemical detection of cancer biomarker interleukin-6 (IL-6) in diluted serum. Capture antibodies of IL-6 were chemically linked onto the electrode arrays and a sandwich immunoassay protocol was developed. A biotinylated detection antibody with polymerized horseradish peroxidase labels was used for signal amplification. The detection limit of IL-6 in diluted serum was remarkably low at 10 fg mL(-1) (385 aM) with a linear response with log of IL-6 concentration from 10 to 1300 fg mL(-1). These easily fabricated, ultrasensitive, microfluidic immunosensors should be readily adapted for sensitive detection of multiple biomarkers for cancer diagnostics.     

Partially fluorinated thermally responsive latices of linear and crosslinked copolymers

A series of thermally responsive copolymers of N-isopropylacrylamide (NIPAAM) with a fluorinated hydrophobic comonomer, either hexafluoroisopropylmethacrylate (HFIPMA) or 2,2,3,3,4,4-hexafluorobutylmethacrylate (HFBMA) and a hydrophilic comonomer, methacrylic acid (MAA), were synthesized by emulsion polymerization. The chemical structures of the copolymers were studied by the IR technique. Dynamic light scattering (DLS) showed that aqueous latices of the copolymers exhibited swelling–deswelling changes typical to PNIPAAM; the degree of swelling as well as the temperature at which the polymers collapse depended on the chemical structure of the comonomers. Endotherms related to the contraction of the polymers were studied by differential scanning calorimetry (DSC). A combination of DLS and DSC results revealed that the hydrophobic and hydrophilic units in the copolymers strongly affected the swelling behavior, as well as the local environment of the PNIPAAM chains. The comonomer HFIPMA increased the hydrophobicity of NIPAAM, reduced the swelling, and caused coagulation of the copolymer of NIPAAM and HFIPMA at temperatures above the critical temperature. Hydrophobicity of HFIPMA also affected the rheological properties of the latex. The HFBMA comonomer increased the swelling of the latex particles. Methacrylic acid added into the associating copolymers made the copolymers to show polyelectrolyte behavior with an increase of swelling and a decrease of the enthalpy change upon the collapse. © 1998 John Wiley & Sons, Inc. J. Polym. Sci. B Polym. Phys. 36: 2141–2152, 1998     

Optical properties of thermally responsive amphiphilic gold nanoparticles protected with polymers

Amphiphilic thermally responsive gold nanoparticles have been prepared by protecting the particles with both polystyrene, PS, and poly(N-isopropylacrylamide), PNIPAM, chains. Particles form a monolayer on a water surface in a Langmuir trough, and according to in situ spectroscopic measurements, the surface plasmon resonance, SPR, band undergoes a blue-shift during the monolayer compression. The compression-induced blue-shift is related to a change in the conformation of tethered PNIPAM chains; the phenomenon is discussed on the basis of Mie-Drude theory. In contrast, a red-shift in the SPR of the multilayers of the same nanoparticles transferred at different temperatures has been observed with increasing the deposition cycle, attributed to the presence of a weak interparticle coupling in the multilayer.     

Fabrication of aligned TiO2 one-dimensional nanostructured arrays using a one-step templating solution approach

In this letter, we report a one-step templating synthetic strategy to prepare aligned TiO2 nanotube and nanowire arrays on Si substrate from a solution at ambient temperature. The deposition of TiO2 and the selective-etching of the ZnO template proceeded at the same time through the careful control of process parameters. The different thickness of TiO2 sheaths, leading to the formation of nanotubes or nanorods, can be precisely controlled by the deposition time. The idea of selective etching and deposition is applicable to other oxide materials, and such a facile method is expected to find widespread applications.     

Simple conjugation and purification of quantum dot-antibody complexes using a thermally responsive elastin-protein L scaffold as immunofluoresecent agents

Fluorescent quantum dots (QDs), because of their tunable spectral properties, are ideal for simultaneous multiplexed detection in an antibody array format. Despite these advantages, their widespread usage is limited by the costly and tedious conjugation and separation protocol. Herein, we report a simple platform for the direct conjugation and separation of highly luminescent CdSe-ZnS QD-antibody complexes using a genetically engineered polyhistidine tagged elastin-protein L fusion (His-ELP-PL). The principle of immunoassay-ready conjugates was to take advantage of the direct conjugation of QDs via metal coordination with the His tag, the unique temperature-responsive property of ELP, and the high affinity of the antibody-binding protein L domain toward IgGs. Simple separation of the QD- His-ELP-PL-IgG complex was achieved by thermally triggered precipitation without any interference on the QD functionality. The utility of the biofunctionalized OD probes was demonstranted in an antibody array for the detection of carcinoembryonic antigen.     

Fabrication and supercapacitive performance of long anodic TiO2 nanotube arrays using constant current anodization

A galvanostatic anodization is used to prepare long TiO₂ nanotube arrays (TNTAs). TNTAs of over 100 μm in length, with similar nanotube size and structural regularity to the classic TNTAs made from potentiostatic mode, are achieved at 10 mA cm^− 2. After a post-anodization in a H₃PO₄-based electrolyte, the TNTAs with long nanotubes exhibit good adhesion to Ti substrate. The as-prepared long TNTAs yield a larger areal capacitance of 128.4 mF cm^− 2. Further, the long TNTAs possess a higher surface area, making them suitable as support templates for other active materials.     

Fabrication of titania nanotube arrays on curved surface

In this paper, well-ordered titania nanotube arrays were formed on curved surface provided by titanium wire via anodic oxidation. The morphology of the nanotube arrays was observed by scanning electron microscopy (SEM). It was found that within the range of 360°, all the growing orientations of each nanotube keep correspondence with their outer electric fields. That is to say the surface shapes of anode play an important role on morphology of nanotube arrays. Compared with foils, by changing the anode shape, higher aspect ratio can be obtained. The nanotube arrays formed on wires can provide good understanding for the formation mechanism of the nanotube arrays. Furthermore, it can stimulate new thoughts in practical applications due to its ringed shaper.     

Fabrication and characterization of single-crystalline ZnTe nanowire arrays

Semiconductor ZnTe nanowire arrays have been synthesized by the pulsed electrochemical deposition from aqueous solutions into porous anodic alumina membranes. X-ray diffraction analyses show that the as-synthesized nanowires have a highly preferential orientation. Scanning electron microscopy, transmission electron microscopy, and high-resolution transmission electron microscopy indicate that high-filling, ordered, and single-crystalline nanowire arrays have been obtained. The optical absorption spectra of the nanowire arrays show that the optical absorption band edge of the ZnTe nanowire array exhibits a blue shift compared with that of bulk ZnTe. The growth mechanism and the electrochemical deposition process are discussed together with the chemical compositions analysis.     

Fabrication and electrochemical characterization of interdigitated nanoelectrode arrays

Interdigitated nanoelectrode arrays with controlled electrode bandwidth and gap geometries ranging from 30 nm to 1 μm were fabricated on glass substrates by a planar process involving high resolution electron beam lithography and lift-off, and their characteristic electrochemical responses to an aqueous ferrocene derivative solution were examined using fundamental electrochemical techniques. Despite the comparatively large electrode area of electrode arrays containing 10 bands to a single band electrode, quasi-steady-state currents with high current density were obtained at a slow potential sweep rate in cyclic voltammograms of ferrocene derivative since the lateral dimension of the nanoelectrode arrays was considerably less than the scale of the diffusion layer of redox species. Additionally, it was demonstrated that the electrode thickness influenced limiting currents of voltammograms in the case of nanoelectrode arrays. In generation-collection mode experiments, furthermore, a collection efficiency as high as ∼99% was attained by 100 nm wide electrode arrays with a gap dimension of 30 nm.     

Organelle-specific zinc detection using zinpyr-labeled fusion proteins in live cells

A protein labeling approach is employed for the localization of a zinc-responsive fluorescent probe in the mitochondria and in the Golgi apparatus of living cells. ZP1, a zinc sensor of the Zinpyr family, was functionalized with a benzylguanine moiety and thus converted into a substrate (ZP1BG) for the human DNA repair enzyme alkylguaninetransferase (AGT or SNAP-Tag). The labeling reaction of purified glutathione S-transferase tagged AGT with ZP1BG and the zinc response of the resulting protein-bound sensor were confirmed in vitro. The new detection system, which combines a protein labeling methodology with a zinc fluorescent sensor, was tested in live HeLa cells expressing AGT in specific locations. The enzyme was genetically fused to site-directing proteins that anchor the probe onto targeted organelles. Localization of the zinc sensors in the Golgi apparatus and in the mitochondria was demonstrated by fluorescence microscopy. The protein-bound fluorescence detection system is zinc-responsive in living cells.     

Simultaneous detection of small molecules,proteins and microRNAs using single molecule arrays

Biological samples such as blood, urine, cerebrospinal fluid and saliva contain a large variety of proteins, nucleic acids, and small molecules. These molecules can serve as potential biomarkers of disease and therefore, it is desirable to simultaneously detect multiple biomarkers in one sample. Current detection techniques suffer from various limitations including low analytical sensitivity and complex sample processing. In this work, we present an ultrasensitive method for simultaneous detection of small molecules, proteins and microRNAs using single molecule arrays (Simoa). Dye-encoded beads modified with specific capture probes were used to quantify each analyte. Multiplex competitive Simoa assays were established for simultaneous detection of cortisol and prostaglandin E2. In addition, competitive and sandwich immunoassays were combined with a direct nucleic acid hybridization assay for simultaneous detection of cortisol, interleukin 6 and microRNA 141. The multi-analyte Simoa assay shows high sensitivity and specificity, which provides a powerful tool for the analysis of many different samples.

The first example of multiplexed detection of proteins, nucleic acids, and small molecules using single molecule measurement methodology.     

Hydrogel-templated growth of large gold nanoparticles: synthesis of thermally responsive hydrogel-nanoparticle composites

In this paper, we describe a unique strategy for preparing discrete composite nanoparticles consisting of a large gold core (60-150 nm in diameter) surrounded by a thermally responsive nontoxic hydrogel polymer derived from the polymerization of N-isopropylacrylamide (NIPAM) or a mixture of NIPAM and acrylic acid. We synthesize these composite nanoparticles at room temperature by inducing the growth of gold nanoparticles in the presence of preformed spherical hydrogel particles. This new method allows precise control of the size of the encapsulated gold cores (tunable between 60 and 150 nm) and affords composite nanoparticles possessing diameters ranging from as small as 200 nm to as large as 550 nm. Variable-temperature studies show that the hydrodynamic diameter of these composite nanoparticles shrinks dramatically when the temperature is increased above the lower critical solution temperature (LCST); correspondingly, when the temperature is lowered below the LCST, the hydrodynamic diameter expands to its original size. These composite nanoparticles are being targeted for use as optically modulated drug-delivery vehicles that undergo volume changes upon exposure to light absorbed by the gold nanoparticle core.     

Steric stabilization of thermally responsive N-isopropylacrylamide particles by poly(vinyl alcohol)

Poly(vinyl alcohol) (PVA) was used as a steric stabilizer for the dispersion polymerization of cross-linked poly(N-isopropylacrylamide) (PNIPAM) in water. A series of reactions were carried out using PVA of varying molecular weight and degree of hydrolysis. Under appropriate conditions, PNIPAM particles of uniform and controllable size were produced using PVA as the stabilizer. The colloidal stability was investigated by measuring changes in particle size with temperature in aqueous suspensions of varying ionic strength. For comparison, parallel colloidal stability measurements were conducted on PNIPAM particles synthesized with low-molecular-weight ionic surfactants. PVA provides colloidal stability over a wide range of temperature and ionic strength, whereas particles produced with ionic surfactants flocculate in moderate ionic strength solutions upon collapse of the hydrogel as the temperature is increased. Experimental results and theoretical consideration indicate that sterically stabilized PNIPAM particles resulted from the grafting of PVA to the PNIPAM particle surface. The enhanced colloidal stability afforded by PVA allows the temperature-responsive PNIPAM particles to be used under physiological conditions where electrostatic stability is ineffective.     

Fabrication of multi-sectional TiO 2 nanotube arrays by anodization

We report a facile method to grow multi-sectional TiO₂ nanotube arrays consisting of alternating bamboo-shaped and smooth-walled nanotube sections by anodization. Two key factors are necessary for obtaining these morphologies. First, in order to avoid possible disruptions between the conjoint sections of the nanotube, the distribution of hydrogen ions is suggested not to be fiercely disturbed when switching from the first to the second stage. Second, to avoid the disruption of the nanotube at the joint which results from the disparity in diameters between sections, the direct current voltage is set to be the maximum of the square wave voltage. These newly developed TiO₂ nanotube arrays are expected to have potential applications in solar cells, drug release and delivery systems.