Capillary array scanner for time-resolved detection and identification of fluorescently labelled DNA fragments

The first capillary array scanner for time-resolved fluorescence detection in parallel capillary electrophoresis based on semiconductor technology is described. The system consists essentially of a confocal fluorescence microscope and a x,y-microscope scanning stage. Fluorescence of the labelled probe molecules was excited using a short-pulse diode laser emitting at 640 nm with a repetition rate of 50 MHz. Using a single filter system the fluorescence decays of different labels were detected by an avalanche photodiode in combination with a PC plug-in card for time-correlated single-photon counting (TCSPC). The time-resolved fluorescence signals were analyzed and identified by a maximum likelihood estimator (MLE). The x,y-microscope scanning stage allows for discontinuous, bidirectional scanning of up to 16 capillaries in an array, resulting in longer fluorescence collection times per capillary compared to scanners working in a continuous mode. Synchronization of the alignment and measurement process were developed to allow for data acquisition without overhead. Detection limits in the subzeptomol range for different dye molecules separated in parallel capillaries have been achieved. In addition, we report on parallel time-resolved detection and separation of more than 400 bases of single base extension DNA fragments in capillary array electrophoresis. Using only semiconductor technology the presented technique represents a low-cost alternative for high throughput DNA sequencing in parallel capillaries.     

SCF-Xα-SW model calculations for metal clusters of nickel,copper, and silver and for the oxygen chemisorption on these metals

The oxygen chemisorption on Ni, Cu and Ag is studied by comparing PE spectra of these systems and SCF-Xα scattered-wave cluster models. Consideration of octahedral clusters M₆ (M = Ni, Cu, Ag) shows that they are large enough to reproduce trends in energy differences, such as the width of the d-bands and the distance from the top of the d-bands to the Fermi level, as found in experiment and in bulk energy band calculations. Substrate model clusters for the interaction of oxygen with different metal surfaces are derived from an octahedron by removing one ((100) face) or two adjacent metal atoms ((110) face). Comparing the UPS difference spectrum for O/Ag (110) with several Ag₄O cluster models makes it possible to interpret the peaks above the Ag d-band as O-Ag anti-bonding levels. These peaks are caused by O 2p-Ag 4d and O 2p-Ag 5s interaction. The corresponding bonding levels fall in the Ag d-bands and cannot therefore be identified with confidence in the spectra. The decreasing intensity of the oxygen derived peak below the metal d-band in the UPS spectra when going from Ni to Cu to Ag, and the simultaneously increasing O peaks above the d-band correlate with the changes of the localization of the corresponding bonding and anti-bonding levels in the oxygen sphere and the decreasing strength of the chemisorption bond.     

Soliton concept in general relativity

Soliton physics has made considerable progress in solving nonlinear problems. This paper is meant to relate the soliten concept to the stationary axisymmetric vacuum fields in general relativity. We present a functional transformation which, working as a nonlinear creation operator, generates gravitational fields of isolated sources. When applied to flat space-time (gravitational vacuum) this operation leads to a nonlinear superposition of an arbitrary number of Kerr particles. This superposition also includes the Tomimatsu-Sato fields. The functional transformations form an infinite-parameter group which contains the Kinnersley-Geroch group as a subgroup.This essay received the second award from the Gravity Research Foundation for the year 1980.-Ed.     

Characterisation of Phosphorus Compounds By 31P,NX-Shift Correlated NMR Techniques in One and Two Dimensions

Abstract

Examples for 2-D shift correlations between ³¹P and another heteronucleus via ⁿX-detected 2D-INJZPT or ³¹P-detected HMQC experiments are presented. The former technique is best suited for ³¹P, ¹⁵N correlations and permits the determination of intramolecular connectivities as well as of relative signs of couplings.¹ The “inverse” HMQC expenment allows easy recording of the ⁷⁷Se NMR spectrum of an equilibrium mixture of 2 and 3³, disregarding the severe exchange broadening of the ⁷⁷Se-resonances     

High Stability of a Phosphenium Ion Allows Umpolung of P--H Bonds

The high intrinsic stability of 1,3,2-diazaphospholenium cations enhances ionic polarization of covalent P--X bonds in P -halogeno- and P -hydrido-diazaphospholenes. The physical properties of the latter suggest a hydridic nature of the P--H bond, and their reactivities display an "Umpolung" as compared to known reaction patterns of phosphines.     

Screening Dyrk1A inhibitors by MALDI-QqQ mass spectrometry: systematic comparison to established radiometric,luminescence, and LC–UV–MS assays

Enzyme-catalyzed reactions play key roles in disease pathology, thus making them relevant subjects of therapeutic inhibitor screening experiments. Matrix-assisted laser desorption/ionization (MALDI) assays have been demonstrated to be able to replace established screening approaches. They offer increased sample throughput, but care must be taken to avoid instrumental bias from differences in ionization efficiencies. We compared a MALDI-triple-quadrupole (QqQ) method for the Dyrk1A peptide substrate woodtide to LC–MS, liquid chromatography with ultraviolet detection (LC–UV), luminescence, and radiometric assays. MALDI measurements were performed on a MALDI-QqQ instrument in the multiple-reaction monitoring mode. Different MALDI conditions were investigated to address whether matrix type, sample support, and MRM- or SIM-based detection conditions can be used to accommodate the molar responses of substrate peptide and its phosphorylated form. UV detection served as a reference method. The impact of MALDI matrix on IC₅₀ values was small, even considering that matrix preparations were used that are known to alleviate response differences. IC₅₀ values determined by MALDI were ca. 2-fold lower than those determined by LC–UV. Although MALDI generated lower ion yields for the phosphorylated peptide than for the peptide substrate, we found that a correction of compound potencies was readily possible using correction factors based on unbiased LC–UV results. A thorough method development delivered a robust assay with excellent performance (Z′ > 0.91) that was close to that seen for LC–UV.

Highly Selective Dimerization and Trimerization of Isobutene to Linearly Linked Products by Using Nickel Catalysts

The unique linear linkage of isobutene to generate highly valuable C₈ precursors for plasticizers is feasible by using special nickel catalysts. (4‐Cyclooctene‐1‐yl)(1,1,1,5,5,5‐hexafluoro‐2,4‐acetylacetonato)nickel and aluminum‐alkyl‐activated nickel acetylacetonates produce isobutene dimers with high selectivities of up to 95 %. Moreover, selectivity for the head‐to‐head products (2,5‐dimethylhexenes) is remarkably high at up to 99 %. Additionally, novel C₁₂ isobutene trimers are also formed with a very high selectivity of up to 99 % for the linear linkage. The trimer structure (2,5,8‐trimethylnonenes) reflects the stepwise characteristic of the reaction mechanism. Pathways of insertion and activation and the deactivation processes of the catalyst are discussed in detail.