Rapid activation of the melibiose permease MelB immobilized on a solid-supported membrane

Rapid solution exchange on a solid-supported membrane (SSM) is investigated using fluidic structures and a solid-supported membrane of 1 mm diameter in wall jet geometry. The flow is analyzed with a new technique based on specific ion interactions with the surface combined with an electrical measurement. The critical parameters affecting the time course of the solution exchange and the transfer function describing the time resolution of the SSM system are determined. The experimental data indicate that solution transport represents an intermediate situation between the plug flow and the Hagen-Poiseuille laminar flow regime. However, to a good approximation the rise of the surface concentration can be described by Hagen-Poiseuille flow with ideal mixing at the surface of the SSM. Using an improved cuvette design, solution exchange as fast as 2 ms was achieved at the surface of a solid-supported membrane. As an application of the technique, the rate constant of a fast electrogenic reaction in the melibiose permease MelB, a bacterial ( Escherichia coli) sugar transporter, is determined. For comparison, the kinetics of a conformational transition of the same transporter was measured using stopped-flow tryptophan fluorescence spectroscopy. The relaxation time constant obtained for the charge displacement agrees with that determined in the stopped-flow experiments. This demonstrates that upon sugar binding MelB undergoes an electrogenic conformational transition with a rate constant of k approximately 250 s (-1).     

Bioactive textiles by sol–gel immobilised natural active agents

The sol–gel immobilisation and controlled release of various bioactive liquids (BL) from within modified silica coatings were investigated, in order to evaluate the suitability of functionalised textiles for the following applications: (1) skin-friendly textiles with antimicrobial and antiallergic effects due to immobilised natural oils such as evening primrose and perilla oil; (2) textiles for therapeutic treatment of the respiratory tract by means of immobilised mixtures of high volatility natural agents such as eucalyptol, camphor and menthol. Results indicating the antimicrobial properties, washfastness and medical activity of the bioactive textiles are presented. Of note is the transdermal resorption of the BL from the sol–gel coated textiles, determined by the time-dependent concentration of cineol, camphor and menthol in the air exhaled by test subjects after the application of coated textiles. When compared to the application of an ointment, not only does the use of coated textiles offer a more continuous and prolonged release of the embedded BL, but it is also more convenient.     

Surface acoustic wave biosensors: a review

This review presents an overview of 20 years of worldwide development in the field of biosensors based on special types of surface acoustic wave (SAW) devices that permit the highly sensitive detection of biorelevant molecules in liquid media (such as water or aqueous buffer solutions). 1987 saw the first approaches, which used either horizontally polarized shear waves (HPSW) in a delay line configuration on lithium tantalate (LiTaO₃) substrates or SAW resonator structures on quartz or LiTaO₃ with periodic mass gratings. The latter are termed “surface transverse waves” (STW), and they have comparatively low attenuation values when operated in liquids. Later Love wave devices were developed, which used a film resonance effect to significantly reduce attenuation. All of these sensor approaches were accompanied by the development of appropriate sensing films. First attempts used simple layers of adsorbed antibodies. Later approaches used various types of covalently bound layers, for example those utilizing intermediate hydrogel layers. Recent approaches involve SAW biosensor devices inserted into compact systems with integrated fluidics for sample handling. To achieve this, the SAW biosensors can be embedded into micromachined polymer housings. Combining these two features will extend the system to create versatile biosensor arrays for generic lab use or for diagnostic purposes. SAW based biosensor immersed in a sample flow. Analyte molecules binding to the immobilized antibodies on the sensor surface will influence the velocity of the SAW and hence the output signal generated by the driving electronics.     

N-K electron energy-loss near-edge structures for TiN/VN layers: an ab initio and experimental study

We study N-K-edge electron energy-loss near-edge structures for well-defined TiN/VN bilayers grown on a MgO(100) substrate by both calculations and experiments. The structural relaxations and the electronic structure of TiN/VN multilayers are calculated using the Vienna Ab Initio Simulation Package computer code, which uses density functional theory to describe the electronic interaction. The effects of the core hole created in the excitation process are included in the calculations. For VN, off-stoichiometric effects due to nitrogen vacancies are modelled. The partial density of states (PDOS) for the N-K edge of atoms in the vicinity of the TiN/MgO interface revealed that two new peaks appear between 7 and 9 eV instead of a broad shoulder typical for the bulk. For the VN/TiN interface, the PDOS is modified only slightly, owing to similar bonding on both sides of the interface, and is thus very similar to the respective bulk spectra. An experimental spectrum taken at the VN/TiN interface is, however, well described by an average of the simulated spectra for VN and TiN bulk (interface). Such a finding is characteristic of an intermixed interface.      

A simplified hollow-fibre supported liquid membrane extraction method for quantification of 2-amino-1-methyl-6-phenylimidazo[4,5-<Emphasis Type="BoldItalic">b</Emphasis>]pyridine (PhIP) in urine and plasma samples

A simple and easy-to-use extraction procedure has been optimised, validated, and applied for extraction of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in urine and spiked plasma samples. PhIP is a carcinogenic and mutagenic heterocyclic aromatic amine that is formed during cooking of meat and fish. The novelty of the extraction procedure lies in using a short piece of narrow capillary-like microporous hollow-fibre (HF) membrane as extraction device. The HF membrane was filled with a few microlitres of acidic solution and the membrane pores were impregnated with an organic extraction solvent. Therefore, the technique was called hollow-fibre supported liquid membrane (HF-SLM) extraction. The HF extraction device was then supported by a syringe needle and directly immersed in urine (1.4 mL) or plasma (0.3 mL) previously made alkaline by adding 0.5 mol L⁻¹ NaOH solution to give a final volume of 1.6 mL. The operation of the HF-SLM extraction at the optimal conditions resulted in a PhIP extraction efficiency of 74% from both spiked urine and plasma, corresponding to enrichment factors of 126 and 27, respectively. For 90 min extraction time, limits of detection and quantification were, respectively, 8 and 25 pg mL⁻¹ for urine and 6 and 11 pg mL⁻¹ for plasma. Within-day repeatability (n = 6) and between-day reproducibility (n = 3) were, respectively, 5% and 13% for urine and 6% and 7% for plasma. Analysis of urine samples collected for 12 h after a volunteer had eaten 250 g well-done chicken showed the PhIP concentration was 124 ± 21 pg mL⁻¹, calculated assuming an extraction efficiency of 74%.     

Assembly of a trinuclear metallo-capsule from a tripodal tris(beta-diketone) derivative and copper(II)

A new metallo-capsule has been synthesised that consists of three copper(II) ions and two molecules of a tris-deprotonated tripodal ligand in which three 2,4-pentanedione groups are linked via their gamma-carbons through thioether spacers to the 1,3,5-positions of a triazine core.     

Development and validation of a sensitive enzyme immunoassay for surveillance of Cry1Ab toxin in bovine blood plasma of cows fed Bt-maize (MON810)

The increasing global adoption of genetically modified (GM) plant derivatives in animal feed has provoked a strong demand for an appropriate detection method to evaluate the existence of transgenic protein in animal tissues and animal by-products derived from GM plant fed animals. A highly specific and sensitive sandwich enzyme immunoassay for the surveillance of transgenic Cry1Ab protein from Bt-maize in the blood plasma of cows fed on Bt-maize was developed and validated according to the criteria of EU-Decision 2002/657/EC. The sandwich assay is based on immuno-affinity purified polyclonal antibody raised against Cry1Ab protein in rabbits. Native and biotinylated forms of this antibody served as capture antibody and detection antibody for the ELISA, respectively. Streptavidin-horseradish peroxidase conjugate and TMB substrate provided the means for enzymatic colour development.The immunoassay allowed Cry1Ab protein determination in bovine blood plasma in an analytical range of 0.4-100 ng mL⁻¹ with a decision limit (CCα) of 1.5 ng mL⁻¹ and detection capability (CCβ) of 2.3 ng mL⁻¹. Recoveries ranged from 89 to 106% (mean value of 98%) in spiked plasma.In total, 20 plasma samples from cows (n = 7) fed non-transgenic maize and 24 samples from cows (n = 8) fed transgenic maize (collected before and, after 1 and 2 months of feeding) were investigated for the presence of the Cry1Ab protein. There was no difference amongst both groups (all the samples were below 1.5 ng mL⁻¹; CCα). No plasma sample was positive for the presence of the Cry1Ab protein at CCα and CCβ of the assay.