Thermostable DNA immobilization and temperature effects on surface hybridization

Monolayer films of nucleic acids on solid supports are encountered in a range of diagnostic and bioanalytical applications. These applications often rely on elevated temperatures to improve performance; moreover, studies at elevated temperatures can provide fundamental information on layer organization and functionality. To support such applications, this study compares thermostability of oligonucleotide monolayers immobilized to gold by first coating the gold with a nanometer-thick film (an "anchor layer") of a polymercaptosiloxane, to which DNA oligonucleotides are subsequently tethered through maleimide-thiol conjugation, with thermostability of monolayers formed via widely used attachment through a terminal thiol moiety on the DNA. The temperature range covered is from 25 to 90 °C. After confirming stability of immobilization and, more importantly, retention of hybridization activity even under the harshest conditions investigated, these thermostable films are used to demonstrate measurements of (1) reversible surface melting transitions and (2) temperature dependence of competitive hybridization, when fully matched and mismatched sequences compete for binding to immobilized DNA oligonucleotides. The competitive hybridization experiments reveal a pronounced impact of temperature on rates of approach to equilibrium, with kinetic freezing into nonequilibrium states close to room temperature and rapid approach to equilibrium at elevated temperatures. Modeling of competitive surface hybridization equilibria using thermodynamic parameters derived from surface melting transitions of the individual sequences is also discussed.     

Evaluation of polyphenolic profile and nutritional value of non-traditional fruit species in the czech republic - a comparative study

Dry matter, organic acids, ascorbic acid, minerals (nitrogen, phosphorus, potassium, calcium, magnesium, sodium) and polyphenolic profile of a number of non-traditional fruit species and their genotypes, namely blue honeysuckle (Lonicera spp.), Saskatoon berry (Amelanchier alnifolia), black mulberry (Morus nigra), Tomentosa cherry (Prunus tomentosa Thunb.) and jostaberry (Ribes nigrum x Grossularia uva-crispa) were investigated. The results showed that Lonicera genotypes displayed high levels of ascorbic acid and they were rich in minerals, with the cultivar 'Amfora' achieving the leading position in nitrogen, phosphorus and potassium content among all lesser known fruit species. Amelanchier cultivars represented a valuable source of ascorbic acid and calcium, 'Ti?ňovsky' and 'Smoky' together with Morus nigra 'Jugoslavska' accumulated the highest level of examined polyphenolic compounds. Regular consumption of studied less common fruit species can bring health benefits so they can represent a high potential value for fruit growers and in addition they can be utilised as functional foods.     

Interpolation theorem for Marcinkiewicz spaces with applications to Lorentz gamma spaces

This paper is devoted to the interpolation principle between spaces of weak type. We characterise interpolation spaces between two Marcinkiewicz spaces in terms of Hardy type operators involving suprema. We study general properties of such operators and their behavior on Lorentz gamma spaces. A particular emphasis is placed on elementary and comprehensive proofs.     

Rubber magnets based on NBR and lithium ferrite with the ability to absorb electromagnetic radiation

Rubber magnetic composites were prepared through the incorporation of magnetic soft lithium ferrite into acrylonitrile butadiene rubber. Standard sulfur‐based curing and peroxide curing systems were used for cross‐linking of rubber matrices. The experimental part was focused on the investigation of ferrite content and curing system composition on cross‐link density, physical‐mechanical, magnetic and shielding characteristics of composites. The results demonstrated that cross‐link density and physical‐mechanical properties of composites can be modified by both the amount of ferrite and composition of the curing system. The influence of curing systems on magnetic properties was negligible. It can be stated that the application of lithium ferrite to rubber matrix leads to the preparation of rubber composites with the ability to efficiently absorb harmful electromagnetic radiation in the tested frequency range. The shielding efficiency of composites increased with increasing content of magnetic filler.     

Kinetic mechanisms in morpholino-DNA surface hybridization

Morpholinos (MOs) are DNA analogues whose uncharged nature can bring fundamental advantages to surface hybridization technologies such as DNA microarrays, by using MOs as the immobilized, or "probe", species. Advancement of MO-based diagnostics, however, is challenged by limited understanding of the surface organization of MO molecules and of how this organization impacts hybridization kinetics and thermodynamics. The present study focuses on hybridization kinetics between monolayers of MO probes and DNA targets as a function of the instantaneous extent of hybridization (i.e., duplex coverage), total probe coverage, and ionic strength. Intriguingly, these experiments reveal distinct kinetic stages, none of which are consistent with Langmuir kinetics. The initial stage, in which duplex coverage remains relatively sparse, indicates confluence of two effects: blockage of target access to unhybridized probes by previously formed duplexes and deactivation of the solid support due to consumption of probe molecules. This interpretation is consistent with a surface organization in which unhybridized MO probes localize near the solid support, underneath a layer of MO-DNA duplexes. As duplex coverage builds, provided saturation is not reached first, the initial stage can transition to an unusual regime characterized by near independence of hybridization rate on duplex coverage, followed by a prolonged approach to equilibrium. The possible origins of these more complex latter behaviors are discussed. Comparison with published data for DNA and peptide nucleic acid (PNA) probes is carried out to look for universal trends in kinetics. This comparison reveals qualitative similarities when comparable surface organization of probes is expected. In addition, MO monolayers are found capable of a broad range of reactivities that span reported values for PNA and DNA probes.     

An Optimized Protocol for the Synthesis of Peptides Containing trans-Cyclooctene and Bicyclononyne Dienophiles as Useful Multifunctional Bioorthogonal Probes

Despite the great advances in solid-phase peptide synthesis (SPPS), the incorporation of certain functional groups into peptide sequences is restricted by the compatibility of the building blocks with conditions used during SPPS. In particular, the introduction of highly reactive groups used in modern bioorthogonal reactions into peptides remains elusive. Here, we present an optimized synthetic protocol enabling installation of two strained dienophiles, trans-cyclooctene (TCO) and bicyclononyne (BCN), into different peptide sequences. The two groups enable fast and modular post-synthetic functionalization of peptides, as we demonstrate in preparation of peptide-peptide and peptide-drug conjugates. Due to the excellent biocompatibility, the click-functionalization of the peptides can be performed directly in live cells. We further show that the introduction of both clickable groups into peptides enables construction of smart, multifunctional probes that can streamline complex chemical biology experiments such as visualization and pull-down of metabolically labeled glycoconjugates. The presented strategy will find utility in construction of peptides for diverse applications, where high reactivity, efficiency and biocompatibility of the modification step is critical.     

Influence of peroxide curing systems on the performance of natural rubber-based magnetic composites

Rubber magnetic composites were prepared by incorporation of strontium ferrite into natural rubber-based continuous matrix. The prepared rubber compounds have model character and besides the rubber and the filler, they contained only ingredients of peroxide curing system, dicumyl peroxide as curing agent, and ethylene glycol dimethacrylate as co-agent. The work was focused on the evaluation of curing system composition and magnetic filler content on curing process, cross-link density, physical–mechanical and magnetic properties of tested materials. The achieved results revealed that the evaluated properties are dependent on the composition of curing system and on the content of ferrite too. Ferrite incorporated in the rubber matrix imparts magnetic properties to the composites considerably. In addition, the improvement of physical–mechanical properties with doping content of ferrite was observed.     

Magnetic and transport properties of granular and Heusler-type glass-coated microwires

We studied magnetic and structural properties of granular Co_xCu_100−x (5<x<40 at%), Cu₆₃Fe₃₇ and Heusler-type Ni₂MnGa glass-coated microwires. We found that the structure of Co–Cu microwires consists of two phases: fcc Cu for all the samples and fcc α-Co present for higher Co content. In the case of low Co content, Co atoms are distributed within the Cu matrix. The quantity and the size of grains strongly depend on the geometry of the microwire. Co–Cu and Fe–Cu microwires exhibited considerable magnetoresistance (MR). For Co_xCu_100−x microwires at x≥30 the anisotropic contribution to MR has been observed. Temperature dependences of magnetization measured without an external magnetic field (ZFC) and in the presence of a field (FC) show considerable difference below 20 K, indicating the presence of small α-Fe or Co grains embedded in the Cu matrix. Annealed Ni₂MnGa microwires showed ferromagnetic behavior with Curie temperature about 330 K and polycrystalline structure with space group I4/mmm and lattice parameters a=3.75 Å and c=6.78 Å.     

Solution Properties of Azidokojatoiron(III) Complexes

The formation of iron(III) complexes with chelating azidokojate anions L⁻ was investigated in aqueous solutions as a function of the pH and the c(Fe³⁺):c(HL) molar ratio. Based on the stability constants, the distribution among the above complexes, [Fe(H₂O)₆]³⁺, and [Fe(H₂O)₅(OH)]²⁺ were calculated in solutions of various compositions. The complexes are redox stable in aqueous solutions both in the dark and in visible laboratory light. Properties of the investigated azidokojic acid and its iron(III) complexes are compared with those required for therapeutic applications as alternative iron chelators.     

Magnetic composites based on natural rubber prepared by using peroxide and sulfur curing system

In this work, rubber magnetic composites were prepared by incorporation of strontium ferrite into elastomeric matrix based on natural rubber. Cross‐linking of the rubber matrix was performed by using sulfur and peroxide curing system. The study was aimed at the investigation of the type of curing system and magnetic filler content on curing process and cross‐link density of prepared materials. Then, the influence of both factors on physical–mechanical and magnetic properties was observed. The obtained results demonstrate that sulfur‐cured composites show better physical–mechanical properties, especially at lower content of strontium ferrite. With increasing amount of ferrite, the differences between the characteristics of both types of composites became less visible, while peroxide‐cured sample with maximum ferrite content showed superior tensile strength in comparison with tensile strength of maximally filled sample cured with sulfur system. The obtained results demonstrate better compatibility between the rubber and the filler when peroxide system was introduced for cross‐linking of the rubber matrix. Copyright © 2014 John Wiley & Sons, Ltd.