A novel enzyme immunoassay based on potentiometric measurement of molecular adsorption events by an extended-gate field-effect transistor sensor

We developed a novel enzyme immunoassay based on a potentiometric measurement of molecular adsorption events by using an extended-gate field-effect transistor (FET) sensor. The adsorbing rate of a thiol compound on a gold surface was found to depend on the concentration of the compound. To construct an electrochemical enzyme immunoassay system by using the sensor, the enzyme chemistry of acetylcholinesterase (AChE) to generate a thiol compound was used and combined with the enzyme-linked immunosorbent assays (ELISA). After the AChE-catalyzed reaction, the amount of the antigen was obtained by detecting the adsorbing rate of the generated thiol compound on the gold electrode using the FET sensor. The measurement stability was also found to improve when a high frequency voltage of 10 kHz or more was superimposed to the reference electrode. The signal corresponding to a range between 1 and 250 pg/mL of Interleukin 1β was obtained by the FET sensor when a voltage of 1 MHz was superimposed onto the reference electrode. The FET sensor based ELISA used in this measurement technique can successfully detect Interleukin 1β at concentrations as low as 1 pg/mL.     

Existing and emerging detection technologies for DNA (Deoxyribonucleic Acid) finger printing, sequencing, bio- and analytical chips: a multidisciplinary development unifying molecular biology, chemical and electronics engineering

The current status and research trends of detection techniques for DNA-based analysis such as DNA finger printing, sequencing, biochips and allied fields are examined. An overview of main detectors is presented vis-à-vis these DNA operations. The biochip method is explained, the role of micro- and nanoelectronic technologies in biochip realization is highlighted, various optical and electrical detection principles employed in biochips are indicated, and the operational mechanisms of these detection devices are described. Although a diversity of biochips for diagnostic and therapeutic applications has been demonstrated in research laboratories worldwide, only some of these chips have entered the clinical market, and more chips are awaiting commercialization. The necessity of tagging is eliminated in refractive-index change based devices, but the basic flaw of indirect nature of most detection methodologies can only be overcome by generic and/or reagentless DNA sensors such as the conductance-based approach and the DNA-single electron transistor (DNA-SET) structure. Devices of the electrical detection-based category are expected to pave the pathway for the next-generation DNA chips. The review provides a comprehensive coverage of the detection technologies for DNA finger printing, sequencing and related techniques, encompassing a variety of methods from the primitive art to the state-of-the-art scenario as well as promising methods for the future.     

Environmental evaluation of single-use and reusable cups

Goal, Scope and Background The objective of the study was to determine the environmental effects of the resuable cup used during a major event (which took place in Barcelona, Universal Forum of Cultures, 2004), compared with a single-use cup of the same composition (polypropylene) but with different physical characteristics such as mass, shape and capacity. Methods To perform the environmental evaluations and the comparison of both types of cups, the SimaPro software developed and marketed by PRé Consultants was used. The environmental evaluation of the reusable cup was compared with that of a single-use cup using the LCA methodology [6]. The functional unit used was: ‘Serving 1000 liters of draught beverages’. The objective of the study was to find the minimum number of cycles the reusable cup has to do so that its environmental impact is smaller than that of the single-use cup. Results and Conclusions Taking into account all the hypotheses put forward, the study drew the conclusion that the minimum number of uses of the reusable cup necessary for it to have a smaller environmental impact than the single-use cup is 10. The contribution of each process taking part in the entire life cycle of the cups was also studied in detail. In the case of the single-use cup, the most important contribution to all the impact categories is due to the production of polypropylene and the fabrication of the cup, except for the heavy metals category where it is due to the management of the waste coming from the cup’s use. In the case of the reusable cup being used 10 times, the contribution to the different impact categories of the waste generated by the cup’s use is negligible compared to the contribution of the fabrication and washing processes. In addition, the washing process is the one which contributes most to the ozone layer depletion, heavy metals and carcinogens categories. As the number of uses of the reusable cup increases, the contribution to all the environmental impact categories decreases. However, this reduction is not as significant for the ozone layer depletion, heavy metals and carcinogens categories. This is due to the washing process and the fact that the electrical consumption associated with it increases with the number of washings and, consequently, of uses. Recommendations and Perspectives From the environmental point of view, the reusable cup must be used at least 10 times to have less impact than the single-use cup. This is mainly due to the higher weight of the reusable cup and, therefore, the greater amount of raw material needed for its fabrication. If the LCA methodology had been introduced during the design of the reusable cup, its weight would have been lower. This modification would have resulted in a reduction of the environmental impact associated with the use of the reusable cup and, consequently, a smaller number of uses would have been necessary to attain the same level of impact as the single-use cup.     

Zeolite-based cataluminescence sensor for the selective detection of acetaldehyde.

The reactions of acetaldehyde with O atoms in the cages of large-pore zeolites have been discovered to result in light emission. The luminescence characteristics of acetaldehyde vapours passing through the surface of chosen zeolites were studied using a cataluminescence-based detection system. To demonstrate the feasibility of the method, the detection of acetaldehyde using catalysts was studied systematically and a linear response of 0.06-31.2 microg/mL acetaldehyde vapour was obtained. Methanol, ethanol, isopropanol, methylbenzene, chloroform, dichlormethane and acetonitrile did not interfere with the determination of acetaldehyde. Acetaldehyde vapour could also be distinguished from some homologous series such as formaldehyde, cinnamaldehyde, glutaraldehyde and benzaldehyde on this catalyst, possibly due to the stereoselectivity of the zeolite and its specific reaction mechanism. Moreover, acetaldehyde was quantified without detectable interference from formaldehyde in four artificial samples. Thus, this kind of cataluminescence-based sensor could be potentially extended to the analysis of volatile organic compounds in air, and the simple and portable properties of cataluminescence-based sensors could also make them beneficial in many areas of analytical science.     

A surface acoustic wave sensor functionalized with a polypyrrole molecularly imprinted polymer for selective dopamine detection

A surface acoustic wave sensor operating at 104 MHz and functionalized with a polypyrrole molecularly imprinted polymer has been designed for selective detection of dopamine (DA). Optimization of pyrrole/DA ratio, polymerization and immersion times permitted to obtain a highly selective sensor, which has a sensitivity of 0.55°/mM (≈550 Hz/mM) and a detection limit of ≈ 10 nM. Morphology and related roughness parameters of molecularly imprinted polymer surfaces, before and after extraction of DA, as well as that of the non imprinted polymer were characterized by atomic force microscopy. The developed chemosensor selectively recognized dopamine over the structurally similar compound 4‐hydroxyphenethylamine (referred as tyramine), or ascorbic acid,which co‐exists with DA in body fluids at a much higher concentration. Selectivity tests were also carried out with dihydroxybenzene, for which an unexpected phase variation of order of 75% of the DA one was observed. Quantum chemical calculations, based on the density functional theory, were carried out to determine the nature of interactions between each analyte and the PPy matrix and the DA imprinted PPy polypyrrole sensing layer in order to account for the important phase variation observed during dihydroxybenzene injection. Copyright © 2015 John Wiley & Sons, Ltd.     

Key steps in type III secretion system (T3SS) towards translocon assembly with potential sensor at plant plasma membrane

Many plant‐ and animal‐pathogenic Gram‐negative bacteria employ the type III secretion system (T3SS) to translocate effector proteins from bacterial cells into the cytosol of eukaryotic host cells. The effector translocation occurs through an integral component of T3SS, the channel‐like translocon, assembled by hydrophilic and hydrophobic proteinaceous translocators in a two‐step process. In the first, hydrophilic translocators localize to the tip of a proteinaceous needle in animal pathogens, or a proteinaceous pilus in plant pathogens, and associate with hydrophobic translocators, which insert into host plasma membranes in the second step. However, the pilus needs to penetrate plant cell walls in advance. All hydrophilic translocators so far identified in plant pathogens are characteristic of harpins: T3SS accessory proteins containing a unitary hydrophilic domain or an additional enzymatic domain. Two‐domain harpins carrying a pectate lyase domain potentially target plant cell walls and facilitate the penetration of the pectin‐rich middle lamella by the bacterial pilus. One‐domain harpins target plant plasma membranes and may play a crucial role in translocon assembly, which may also involve contrapuntal associations of hydrophobic translocators. In all cases, sensory components in the target plasma membrane are indispensable for the membrane recognition of translocators and the functionality of the translocon. The conjectural sensors point to membrane lipids and proteins, and a phosphatidic acid and an aquaporin are able to interact with selected harpin‐type translocators. Interactions between translocators and their sensors at the target plasma membrane are assumed to be critical for translocon assembly.     

A colorimetric chiral sensor based on chiral crown ether for the recognition of the two enantiomers of primary amino alcohols and amines

A new colorimetric chiral sensor material consisting of three different functional sites such as chromophore (2,4-dinitrophenylazophenol dye), binding site (crown ether), and chiral barrier (3,3'-diphenyl-1,1'-binaphthyl group) was prepared and applied to the recognition of the two enantiomers of primary amino alcohols and amines. Among five primary amino alcohols and two primary amines tested, the two enantiomers of phenylalaninol show the highest difference in the absorption maximum wavelength (Δλ(max)=43.5 nm) and in the association constants (K(S)/K(R)=2.51) upon complexation with the colorimetric chiral sensor material and, consequently, the two enantiomers of phenylalaninol were clearly distinguished from each other by the color difference.     

Multimode selective detection of mercury by chiroptical fluorescent sensors based on methionine/cysteine

Two multimode Hg(II) sensors, L‐MethBQA and L‐CysBQA, were obtained by fusing methionine or S‐methyl cysteine, into a bis‐quinolyl amine‐based chiral podand scaffold. Quinolyl groups serve as the fluorophore and possess nitrogen lone pairs capable of chelating metal ions. On exposure to Hg²⁺ or Zn²⁺, these sensors show signal enhancement in fluorescence. However, Cu²⁺ quenches their fluorescence in 30:70 acetontrile/water. L‐CysBQA complexes with Hg²⁺, producing an exciton‐coupled circular dichroism spectrum with the opposite sign to the one that is produced by Cu²⁺ or Zn²⁺ complexation. L‐CysBQA binds Hg²⁺ more strongly than Zn²⁺ and is shown to differentiate Hg²⁺ from other metal ions, such as Zn²⁺, Cu²⁺, Ni²⁺, and Pb²⁺, exceptionally well. The synergistic use of relatively soft sulfur, quinoline‐based chiral ligands and chiroptically enhanced fluorescence detection results in high sensitivity and selectivity for Hg²⁺. Chirality, 2011. © 2011 Wiley‐Liss, Inc.     

A genetically encoded, high-signal-to-noise maltose sensor

We describe the generation of a family of high-signal-to-noise single-wavelength genetically encoded indicators for maltose. This was achieved by insertion of circularly permuted fluorescent proteins into a bacterial periplasmic binding protein (PBP), Escherichia coli maltodextrin-binding protein, resulting in a four-color family of maltose indicators. The sensors were iteratively optimized to have sufficient brightness and maltose-dependent fluorescence increases for imaging, under both one- and two-photon illumination. We demonstrate that maltose affinity of the sensors can be tuned in a fashion largely independent of the fluorescent readout mechanism. Using literature mutations, the binding specificity could be altered to moderate sucrose preference, but with a significant loss of affinity. We use the soluble sensors in individual E. coli bacteria to observe rapid maltose transport across the plasma membrane, and membrane fusion versions of the sensors on mammalian cells to visualize the addition of maltose to extracellular media. The PBP superfamily includes scaffolds specific for a number of analytes whose visualization would be critical to the reverse engineering of complex systems such as neural networks, biosynthetic pathways, and signal transduction cascades. We expect the methodology outlined here to be useful in the development of indicators for many such analytes.