Immobilization of DNA onto poly(dimethylsiloxane) surfaces and application to a microelectrochemical enzyme-amplified DNA hybridization assay

This paper describes immobilization of DNA onto the interior walls of poly(dimethylsiloxane) (PDMS) microsystems and its application to an enzyme-amplified electrochemical DNA assay. DNA immobilization was carried out by silanization of the PDMS surface with 3-mercaptopropyltrimethoxysilane to yield a thiol-terminated surface. 5'-acrylamide-modified DNA reacts with the pendant thiol groups to yield DNA-modified PDMS. Surface-immobilized DNA oligos serve as capture probes for target DNA. Biotin-labeled target DNA hybridizes to the PDMS-immobilized capture DNA, and subsequent introduction of alkaline phosphatase (AP) conjugated to streptavidin results in attachment of the enzyme to hybridized DNA. Electrochemical detection of DNA hybridization benefits from enzyme amplification. Specifically, AP converts electroinactive p-aminophenyl phosphate to electroactive p-aminophenol, which is detected using an indium tin oxide interdigitated array (IDA) electrode. The IDA electrode eliminates the need for a reference electrode and provides a steady-state current that is related to the concentration of hybridized DNA. At present, the limit of detection of the DNA target is 1 nM in a volume of 20 nL, which corresponds to 20 attomoles of DNA.     

Study of scanning tunneling microscopy images and probable relaxations of the SrTiO3(100) surface by electronic structure calculations

Partial electron density plots were calculated for a model SrTiO₃(100) surface with √5 × √5 ordered oxygen vacancy to examine why the bright spots of the scanning tunneling microscopy (STM) images of SrTiO₃(100) observed in ultrahigh vacuum (UHV) correspond to the oxygen vacancy sites. Possible dependence of the image on the polarity and magnitude of the bias voltage was also discussed on the basis of partial electron density plot calculations. Our study strongly suggests that the UHV STM imaging involves the lowest-lying d-block level of every two Ti³⁺ centers adjacent to an oxygen vacancy, the tip-sample distance involved in the UHV STM experiments is substantially larger than that involved in typical ambient-condition STM imaging, and the Ti⁴⁺ and Ti³⁺ sites of SrTiO₃(100) are reconstructed.     

Combining reverse osmosis and pulsed electrical current electrodialysis for improved recovery of dissolved organic matter from seawater

Dissolved organic carbon (DOC) in the oceans is one of the largest dynamic reservoirs of carbon on earth, comparable in size to the atmospheric reservoir of carbon (as CO₂) in the atmosphere, or to the amount of carbon in all terrestrial and aquatic biota. The concerted efforts of earth scientists, atmospheric scientists, and biologists who study global biogeochemical cycles and the earth's climate have yielded a rather detailed understanding of carbon in the atmosphere and in biota. Marine dissolved organic matter (DOM) is far less well characterized, principally because it exists as a highly diluted mixture of perhaps millions of organic compounds in a highly saline aqueous solution. Prior to 2007, only around 1/3 of marine DOM was typically recovered from seawater for research purposes, regardless of the method of isolation. In 2007, reverse osmosis (RO) and electrodialysis (ED) were coupled to achieve recoveries of 64–93% of marine DOM. The level of residual salts in the concentrated samples, however, still precluded the characterization of marine DOM by solid-state NMR, mass spectrometry, or even elemental analysis. This paper describes a major improvement to the RO/ED method, in which pulsed ED is used (at sea) to reach roughly 100-fold greater removal of salts compared to non-pulsed ED while maintaining comparable recoveries of DOM.     

A weight-function approach for determining watershed initial conditions for combustion waves

In many generic combustion models, one finds that a combustionwave will develop with a specific wave speed. However, thereare possible initial temperature profiles which do not evolveinto such waves, but rather die out to the ambient temperature.There can exist, in some models, a clear distinction betweenthose initial conditions that do evolve into combustion wavesand those that do not; this is sometimes referred to as thewatershed initial condition. When fuel consumption is consideredto be negligible, analytical methods can be used to obtain theexact watershed. In this paper, we consider the problem of determiningpseudo-watersheds and ascertaining the relationship betweenthese pseudo-watersheds and the exact watersheds. In the processa novel weight-function approach for infinite spatial domainsis developed.     

Involvement of membrane sulfhydryls in the activation and maintenance of nutrient transport in chick embryo fibroblasts

At 5 microgram/ml, insulin stimulates hexose, A-system amino acid, and nucleoside transport by serum-starved chick embryo fibroblasts (CEF). This stimulation, although variable, is comparable to that induced by 4% serum. The sulfhydryl oxidants diamide (1-20 micrometer). hydrogen peroxide (500 micrometer), and methylene blue (50 micrometer) mimic the effect of insulin in CEF. PCMB-S,1 a sulfhydryl-reacting compound which penetrates the membrane slowly, has a complex effect on nutrient transport in serum- and glucose-starved CEF. Hexose uptake is inhibited by 0.1-1 mM PCMB-S in a time- and concentration-dependent manner, whereas A-system amino acid transport is inhibited maximally within 10 min of incubation and approaches control rates after 60 min. A differential sensitivity of CEF transport systems is also seen in cells exposed to membrane-impermeant glutathione-maleimide I, designated GS-Mal. At 2 mM GS-Mal reduces the rate of hexose uptake 80-100% in serum- and glucose-starved CEF; in contrast A-system amino acid uptake is unaffected. D-glucose, but not -L-glucose or cytochalasin B, protects against GS-Mal inhibition. These results are consistent with the hypothesis that sulfhydryl groups are involved in nutrient transport and that those sulfhydryls associated with the hexose transport system and essential for its function are located near the exofacial surface of the membrane in CEF.     

Recording long-term potentiation of synaptic transmission by three-dimensional multi-electrode arrays

Background  

Multi-electrode arrays (MEAs) have become popular tools for recording spontaneous and evoked electrical activity of excitable tissues. The majority of previous studies of synaptic transmission in brain slices employed MEAs with planar electrodes that had limited ability to detect signals coming from deeper, healthier layers of the slice. To overcome this limitation, we used three-dimensional (3D) MEAs with tip-shaped electrodes to probe plasticity of field excitatory synaptic potentials (fEPSPs) in the CA1 area of hippocampal slices of 129S5/SvEvBrd and C57BL/6J-Tyr^C-Brd mice.     

High-precision frequency measurements: indispensable tools at the core of the molecular-level analysis of complex systems

This perspective article provides an assessment of the state-of-the-art in the molecular-resolution analysis of complex organic materials. These materials can be divided into biomolecules in complex mixtures (which are amenable to successful separation into unambiguously defined molecular fractions) and complex nonrepetitive materials (which cannot be purified in the conventional sense because they are even more intricate). Molecular-level analyses of these complex systems critically depend on the integrated use of high-performance separation, high-resolution organic structural spectroscopy and mathematical data treatment. At present, only high-precision frequency-derived data exhibit sufficient resolution to overcome the otherwise common and detrimental effects of intrinsic averaging, which deteriorate spectral resolution to the degree of bulk-level rather than molecular-resolution analysis. High-precision frequency measurements are integral to the two most influential organic structural spectroscopic methods for the investigation of complex materials—NMR spectroscopy (which provides unsurpassed detail on close-range molecular order) and FTICR mass spectrometry (which provides unrivalled resolution)—and they can be translated into isotope-specific molecular-resolution data of unprecedented significance and richness. The quality of this standalone de novo molecular-level resolution data is of unparalleled mechanistic relevance and is sufficient to fundamentally advance our understanding of the structures and functions of complex biomolecular mixtures and nonrepetitive complex materials, such as natural organic matter (NOM), aerosols, and soil, plant and microbial extracts, all of which are currently poorly amenable to meaningful target analysis. The discrete analytical volumetric pixel space that is presently available to describe complex systems (defined by NMR, FT mass spectrometry and separation technologies) is in the range of 10^8–14 voxels, and is therefore capable of providing the necessary detail for a meaningful molecular-level analysis of very complex mixtures. Nonrepetitive complex materials exhibit mass spectral signatures in which the signal intensity often follows the number of chemically feasible isomers. This suggests that even the most strongly resolved FTICR mass spectra of complex materials represent simplified (e.g. isomer-filtered) projections of structural space.     

Search for the Decay K L0-->pi0nu nu[over

We performed a search for the K L0-->pi0nu nu[over] decay at the KEK 12-GeV proton synchrotron. No candidate events were observed. An upper limit on the branching ratio for the decay was set to be 6.7 x 10(-8) at the 90% confidence level.