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Hydrogen peroxide (H₂O₂), a strong oxidizer, is a commonly used sterilization agent employed during aseptic food processing and medical applications. To assess the sterilization efficiency with H₂O₂, bacterial spores are common microbial systems due to their remarkable robustness against a wide variety of decontamination strategies. Despite their widespread use, there is, however, only little information about the detailed time-resolved mechanism underlying the oxidative spore death by H₂O₂. In this work, we investigate chemical and morphological changes of individual Bacillus atrophaeus spores undergoing oxidative damage using optical sensing with trapping Raman microscopy in real-time. The time-resolved experiments reveal that spore death involves two distinct phases: (i) an initial phase dominated by the fast release of dipicolinic acid (DPA), a major spore biomarker, which indicates the rupture of the spore’s core; and (ii) the oxidation of the remaining spore material resulting in the subsequent fragmentation of the spores’ coat. Simultaneous observation of the spore morphology by optical microscopy corroborates these mechanisms. The dependence of the onset of DPA release and the time constant of spore fragmentation on H₂O₂ shows that the formation of reactive oxygen species from H₂O₂ is the rate-limiting factor of oxidative spore death.
A field-effect biosensor employing tobacco mosaic virus (TMV) particles as scaffolds for enzyme immobilization is presented. Nanotubular TMV scaffolds allow a dense immobilization of precisely positioned enzymes with retained activity. To demonstrate feasibility of this new strategy, a penicillin sensor has been developed by coupling a penicillinase with virus particles as a model system. The developed field-effect penicillin biosensor consists of an Al-p-Si-SiO₂-Ta₂O₅-TMV structure and has been electrochemically characterized in buffer solutions containing different concentrations of penicillin G. In addition, the morphology of the biosensor surface with virus particles was characterized by scanning electron microscopy and atomic force microscopy methods. The sensors possessed a high penicillin sensitivity of ~ 92 mV/dec in a nearly-linear range from 0.1 mM to 10 mM, and a low detection limit of about 50 µM. The long-term stability of the penicillin biosensor was periodically tested over a time period of about one year without any significant loss of sensitivity. The biosensor has also been successfully applied for penicillin detection in bovine milk samples.
An array of electrically isolated nanoplate field-effect silicon-on-insulator (SOI) capacitors as a new transducer structure for multiparameter (bio-)chemical sensing is presented. The proposed approach allows addressable biasing and electrical readout of multiple nanoplate field-effect capacitive (bio-)chemical sensors on the same SOI chip, as well as differential-mode measurements. The realized sensor chip has been applied for pH and penicillin concentration measurements, electrical monitoring of polyelectrolyte multilayer formation, and the label-free electrical detection of consecutive deoxyribonucleic acid (DNA) hybridization and denaturation events.
High-k perovskite oxide of barium strontium titanate (BST) represents a very attractive multi-functional transducer material for the development of (bio-)chemical sensors. In this work, a Si-based sensor chip containing Pt interdigitated electrodes covered with a thin BST layer (485 nm) has been developed for multi-parameter chemical sensing. The chip has been applied for the contactless measurement of the electrolyte conductivity, the detection of adsorbed charged macromolecules (positively charged polyelectrolytes of polyethylenimine) and the concentration of hydrogen peroxide (H2O2) vapor. The experimental results of functional testing of individual sensors are presented. The mechanism of the BST sensitivity to charged polyelectrolytes and H2O2 vapor has been proposed and discussed.
This work introduces a novel method for the detection of H₂O₂ vapor/aerosol of low concentrations, which is mainly applied in the sterilization of equipment in medical industry. Interdigitated electrode (IDE) structures have been fabricated by means of microfabrication techniques. A differential setup of IDEs was prepared, containing an active sensor element (active IDE) and a passive sensor element (passive IDE), where the former was immobilized with an enzymatic membrane of horseradish peroxidase that is selective towards H₂O₂. Changes in the IDEs’ capacitance values (active sensor element versus passive sensor element) under H₂O₂ vapor/aerosol atmosphere proved the detection in the concentration range up to 630 ppm with a fast response time (<60 s). The influence of relative humidity was also tested with regard to the sensor signal, showing no cross-sensitivity. The repeatability assessment of the IDE biosensors confirmed their stable capacitive signal in eight subsequent cycles of exposure to H₂O₂ vapor/aerosol. Room-temperature detection of H₂O₂ vapor/aerosol with such miniaturized biosensors will allow a future three-dimensional, flexible mapping of aseptic chambers and help to evaluate sterilization assurance in medical industry.
In this study, we show that synthetic sapphire (Al₂O₃), an established implant material, can also serve as a platform material for biosensors comparable to nanocrystalline diamond. Sapphire chips, beads, and powder were first modified with (3-aminopropyl) triethoxysilane (APTES), followed by succinic anhydride (SA), and finally single-stranded probe DNA was EDC coupled to the functionalized layer. The presence of the APTES-SA layer on sapphire powders was confirmed by thermogravimetric analyis and Fourier-transform infrared spectroscopy. Using planar sapphire chips as substrates and X-ray photoelectron spectroscopy (XPS) as surface-sensitive tool, the sequence of individual layers was analyzed with respect to their chemical state, enabling the quantification of areal densities of the involved molecular units. Fluorescence microscopy was used to demonstrate the hybridization of fluorescently tagged target DNA to the probe DNA, including denaturation- and re-hybridization experiments. Due to its high thermal conductivity, synthetic sapphire is especially suitable as a chip material for the heat-transfer method, which was employed to distinguish complementary- and non-complementary DNA duplexes containing single-nucleotide polymorphisms. These results indicate that it is possible to detect mutations electronically with a chemically resilient and electrically insulating chip material.