Fabrication of material-independent morphology gradients for high-throughput applications

Gradient surfaces allow rapid, high-throughput investigations and systematic studies in many disparate fields, including biology, tribology and adhesion. We describe a novel method for the fabrication of material-independent morphology gradients, involving a two-step process of particle erosion followed by a chemical polishing procedure that preferentially removes features with a small radius of curvature as a function of time. Gradients are fabricated on aluminium surfaces, but they may be readily transferred to other materials via a replication technique, which allows for the production of identical roughness gradient samples with any chosen surface chemistry. The gradients have been characterized by means of scanning electron microscopy and optical profilometry. Standard roughness parameters (Ra, Rq, Rz, Sm and Sk) were calculated from optical profilometry data. The roughness has also been assessed over different wavelength windows by means of a fast Fourier transformation approach.     

Differential effects of synchronous and asynchronous multifinger coactivation on human tactile performance

Background  

Repeated execution of a tactile task enhances task performance. In the present study we sought to improve tactile performance with unattended activation-based learning processes (i.e., focused stimulation of dermal receptors evoking neural coactivation (CA)). Previous studies show that the application of CA to a single finger reduced the stationary two-point discrimination threshold and significantly increased tactile acuity. These changes were accompanied by an expansion of the cortical finger representation in primary somatosensory cortex (SI). Here we investigated the effect of different types of multifinger CA on the tactile performance of each finger of the right hand.     

A novel phosphoramidite method for automated synthesis of oligonucleotides on glass supports for biosensor development

Two protocols for functionalization of glass supports with hexaethylene glycol (HEG)-linked oligonucleotides were developed. The first method (standard amidite protocol) made use of the 2-cyanoethyl-phosphoramidite derivative of 4,4′-dimethoxytrityl-protected HEG. This was first coupled to the support by standard solid-phase phosphoramidite chemistry followed by extension with a thymidylic acid icosanucleotide. Stepwise addition of the linker phosphoramidite graduated at 1% (relative to the total sites available) perstep at 50°C resulted in an optimal yield of immobilized oligonucleotides at a density of 2.24 × 10¹⁰ strands/mm². This observed loading maximum lies well below the theoretical maximum loading owing to nonspecific adsorption of HEG on the glass and subsequent blocking of reactive sites. Surface loadings as high as 3.73 × 10¹⁰/mm² and of excellent sequence quality were achieved with a reverse amidite protocol. The support was first modified into a 2-cyanoethyl-N,N-diisopropylphosphoramidite analog followed by coupling with 4,4′-dimethoxytrityl-protected HEG. This protocol is conveniently available when using a conventional DNA synthesizer. The reverse amidite protocol allowed for control of the surface loading at values suitable for subsequent analytical applications that make use of immobilized oligonucleotides as probes for selective hybridization of sample nucleic acids of unknown sequence and concentration.     

[(Mes3Sn)2MoO4], ein monomeres Triorganozinnmolybdat

[(Mes₃Sn)₂MoO₄], a Monomeric Triorganotin Molybdate Mes₃SnBr (Mes = 1, 3, 5‐trimethylphenyl) reacts with (NBu₄)₂[Mo₆O₁₉] in the presence of (NBu₄)OH (in CH₃CN as solvent) to form [(Mes₃Sn)₂MoO₄]. Alternatively the title compound can be obtained from the reaction of [MoO₂(acac)₂] (acac = 2, 4‐pentadionate) with Mes₃SnOH in isopropanol. [(Mes₃Sn)₂MoO₄] forms monoclinic crystals, space group C2/c, with a = 2271.6(3) pm, b = 825.2(1) pm, c = 2739.9(5) pm, β = 90.96(2)°. The crystal structure consists of isolated molecules in which a tetrahedral MoO₄ unit is connected to two terminal Mes₃Sn groups. The Mo‐O distances range from 169.6(4) to 181.1(3) pm and the Sn‐O distance is 204.8(3) pm.     

Rapid quantitative capillary zone electrophoresis method for monitoring the micro-heterogeneity of an intact recombinant glycoprotein

A simple high-resolution capillary zone electrophoresis (CZE) method capable of rapidly assessing the micro-heterogeneity of a 24 kDa molecular weight glycoprotein, has been developed. Separation is carried out using a bare silica capillary at a pH of 2.5 in a commercially available electrophoresis buffer system composed of triethanolamine and phosphoric acid. Over 30 peaks were detected within a run time of 15 min using a 27 cm capillary and approximately 60 peaks were detected using a 77 cm capillary. Although most of the peaks arise from differences in the oligosaccharide structures present on the one glycosylation site on this molecule, other forms of micro-heterogeneity due to the presence of the nonglycosylated form of this glycoprotein and various types of chemical degradation, e.g., deamidation, are also responsible for the multitude of peaks observed. Although the exact chemical identity of each peak in the resulting electropherogram of this glycoprotein is not known, useful information can be obtained for assessing comparability, stability, and batch consistency. Factors impacting the resolution, precision, accuracy, and robustness of the assay are also discussed along with inherent advantages and limitations associated with measuring the micro-heterogeneity of intact glycoproteins.     

Using ultrafast infrared multidimensional correlation spectroscopy to aid in vibrational spectral peak assignments

Ultrafast infrared heterodyne detected vibrational stimulated echoes with full phase information are used to obtain the vibrational correlation spectrum from a mixture of metal-carbonyl compounds. The linear absorption spectrum displays four peaks in the carbonyl stretching region. In the absence of knowledge of the molecules that make up the mixture, the absorption spectrum could arise from four molecules that each produces a single peak to one molecule with four peaks. In contrast, the correlation spectrum displays four peaks on the diagonal and off-diagonal peaks that make it straightforward to determine which peaks belong to a particular molecule.     

Evaluation of multidimensional (ion-exchange/reversed-phase) protein separations using linear and step gradients in the first dimension

The performance characteristics of multidimensional liquid chromatographic protein separations were evaluated using on-line electrospray mass detection, and a novel workflow for automated LC/MS data processing. Two-dimensional ion exchange/reversed-phase LC separations of Escherichia coli cytosol were conducted using either a continuous linear or discontinuous step gradient in the first dimension. Chromatographic profiles of the top 100 most abundant components were characterized to assess overall separation reproducibility within each mode, and to characterize differences in component distribution between the two modes of operation. Analysis of the resulting data indicates that multidimensional separations of complex protein mixtures can be done reproducibly. Furthermore, under the conditions employed within this study, a linear first dimension gradient was more effective at fractionating the protein mixture, distributing fewer major components to multiple second dimension cycles than an equivalent step gradient. The application of on line mass spectrometry, and automated processing of the resulting data, proved valuable for producing component level analysis of multidimensional protein separations.