Development of a Fluorescent Enzyme-Linked DNA Aptamer-Magnetic Bead Sandwich Assay and Portable Fluorometer for Sensitive and Rapid Leishmania Detection in Sandflies

A fluorescent peroxidase-linked DNA aptamer-magnetic bead sandwich assay is described which detects as little as 100 ng of soluble protein extracted from Leishmania major promastigotes with a high molarity chaotropic salt. Lessons learned during development of the assay are described and elucidate the pros and cons of using fluorescent dyes or nanoparticles and quantum dots versus a more consistent peroxidase-linked Amplex Ultra Red (AUR; similar to resazurin) fluorescence version of the assay. While all versions of the assays were highly sensitive, the AUR-based version exhibited lower variability between tests. We hypothesize that the AUR version of this assay is more consistent, especially at low analyte levels, because the fluorescent product of AUR is liberated into bulk solution and readily detectable while fluorophores attached to the reporter aptamer might occasionally be hidden behind magnetic beads near the detection limit. Conversely, fluorophores could be quenched by nearby beads or other proximal fluorophores on the high end of analyte concentration, if packed into a small area after magnetic collection when an enzyme-linked system is not used. A highly portable and rechargeable battery-operated fluorometer with on board computer and color touchscreen is also described which can be used for rapid (<1 h) and sensitive detection of Leishmania promastigote protein extracts (～100 ng per sample) in buffer or sandfly homogenates for mapping of L. major parasite geographic distributions in wild sandfly populations.     

Synthesis of (3E)-Dodecen-12-olide,a Potential Pheromone Component of the Emerald Ash Borer

Emerald ash borer (EAB), Agrilus planipennis Fairmaire, is an invasive insect that has killed millions of ash trees in the USA and Canada. A concise synthesis of a potential EAB pheromone component, (3E)-dodecen-12-olide, using highly stereoselective Julia–Kocienski olefination as the key step, is reported.

Supplemental materials are available for this article. Go to the publisher's online edition of Synthetic Communications® to view the free supplemental file.     

Age-related changes of anisotropic properties of water self-diffusion in human lens: Diffusion anisotropy in lens with aging

Water self-diffusion in human lenses of different age is studied. It is shown that there is a strong anisotropy of self-diffusion of water molecules in old lenses, whereas in young lenses such anisotropy does not occur. This anisotropy appears between 20 and 50 years and is probably connected with the flatness of lens fibers with aging.     

Methods for the rapid solution of the pricing PIDEs in exponential and Merton models

The pricing equations for options on assets that follow jump-diffusion processes contain integrals in addition to the usual differential terms. These integrals usually make such equations expensive to solve numerically. Although Fast Fourier Transform methods can be used to to evaluate the integrals at all mesh points simultaneously, they are costly since the computational region must be extended in order to avoid problems with wrap around. Other numerical difficulties arise when the density function for the jump size is not smooth, as in the Kou double exponential model. We present new solution methods which are based on the fact that even when the problems contain time-dependent parameters the integrals often satisfy easily solved ordinary or parabolic partial differential equations. In particular, we show that by using the operator splitting method proposed by Andersen and Andreasen it is possible to reduce the solution of the pricing equation in the Kou and similar models to a sequence of ordinary differential equations at each time step. We discuss the methods and present results of numerical experiments.     

Dramatic performance enhancements for the FASTER optimization algorithm

FASTER is a combinatorial optimization algorithm useful for finding low-energy side-chain configurations in side-chain placement and protein design calculations. We present two simple enhancements to FASTER that together improve the computational efficiency of these calculations by as much as two orders of magnitude with no loss of accuracy. Our results highlight the importance of choosing appropriate initial configurations, and show that efficiency can be improved by stringently limiting the number of positions that are allowed to relax in response to a perturbation. The changes we describe improve the quality of solutions found for large-scale designs, and allow them to be found in hours rather than days. The improved FASTER algorithm finds low-energy solutions more efficiently than common optimization schemes based on the dead-end elimination theorem and Monte Carlo. These advances have prompted investigations into new methods for force field parameterization and multiple state design.     

A search algorithm for fixed-composition protein design

We present a computational protein design algorithm for finding low-energy sequences of fixed amino acid composition. The search algorithms used in protein design typically do not restrict amino acid composition. However, the random energy model of Shakhnovich suggests that the use of fixed-composition sequences may circumvent defects in the modeling of the denatured state. Our algorithm, FC_FASTER, links fixed-composition versions of Monte Carlo and the FASTER algorithm. As proof of principle, FC_FASTER was tested on an experimentally validated, full-sequence design of the beta1 domain of protein G. For the wild-type composition, FC_FASTER found a lower energy sequence than the experimentally validated sequence. Also, for a different composition, FC_FASTER found the hypothetical lowest-energy sequence in 14 out of 32 trials.     

Application of the multiple isotope material basis set (MIMBS) method of isotope identification to low energy gamma emitters

The multiple isotope material basis set (MIMBS) method for isotope identification combines the material basis set (MBS) model of gamma spectrum attenuation with ordinary response function fitting to identify shielded gamma-emitting isotopes, using low and medium resolution gamma detectors such as NaI and LaBr₃. Although MIMBS has been shown to outperform conventional isotope identification algorithms that do not correct for attenuation effects, it has difficulty identifying low energy emitters such as ⁵⁷Co or ²⁴¹Am. In this article we examine the use of optimized multiple attenuator thicknesses in generating basis spectra for each isotope to obtain better modeling of the low energy spectrum while simultaneously extending the range of the model to thicker attenuators. The effectiveness of the multiple thickness MIMBS algorithm in improving isotope identification rates compared with the original MIMBS method is demonstrated with analyses of simulated gamma spectra. The identification rates obtained with the MIMBS methods are compared to those obtained using the commercial peak-based ScintiVision NaI analysis software.     

A new abietane and two dimeric abietane diterpenes from the black heartwood of Cryptomeria japonica

Three new diterpenes, sugikurojins A-C (1-3) were isolated from the black heartwood of Cryptomeria japonica. The structure of sugikurojin A was deduced to be 19-hydroxy-6,7-dehydroferruginol on the basis of extensive NMR experiments. Sugikurojin B was a dimer of 6,7-dihydroxyferruginol and 6,7-dehydroferruginol with a 6-O-11' linkage. Sugikurojin C was a dimeric ferruginol with 6-O-7' and 7-O-6' linkages. Also obtained in this investigation were the known compounds formosaninol (4), 15 sesquiterpenes (5-19), 16 diterpenes (20-35), three phenylpropanoids (36-38), and a phenolic compound (39).