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Capacitive field-effect electrolyte-insulator-semiconductor sensors consisting of an Al-p-Si-SiO2 structure have been used for the electrical detection of unlabelled single- and double-stranded DNA (dsDNA) molecules by their intrinsic charge. A simple functionalization protocol based on the layer-by-layer (LbL) technique was used to prepare a weak polyelectrolyte/probe-DNA bilayer, followed by the hybridization with complementary target DNA molecules. Due to the flat orientation of the LbL-adsorbed DNA molecules, a high sensor signal has been achieved. In addition, direct label-free detection of in-solution hybridized dsDNA molecules has been studied.
A concept for a new generation of an integrated multi-functional biosensor/actuator system is developed, which is based on biomolecular logic principles. Such a system is expected to be able to detect multiple biochemical input signals simultaneously and in real-time and convert them into electrical output signals with logical operations such as OR, AND, etc. The system can be designed as a closed-loop drug release device triggered by an enzyme logic gate, while the release of the drug induced by the actuator at the required dosage and timing will be controlled by an additional drug sensor. Thus, the system could help to make an accurate and specific diagnosis. The presented concept is exemplarily demonstrated by using an enzyme logic gate based on a glucose/glucose oxidase system, a temperature-responsive hydrogel mimicking the actuator function and an insulin (drug) sensor. In this work, the results of functional testing of individual amperometric glucose and insulin sensors as well as an impedimetric sensor for the detection of the hydrogel swelling/shrinking are presented.
Miniaturized setup, compatibility with advanced micro- and nanotechnologies, and ability to detect biomolecules by their intrinsic molecular charge favor the semiconductor field-effect platform as one of the most attractive approaches for the development of label-free DNA chips. In this work, a capacitive field-effect EIS (electrolyte–insulator–semiconductor) sensor covered with a layer-by-layer prepared, positively charged weak polyelectrolyte layer of PAH (poly(allylamine hydrochloride)) was used for the label-free electrical detection of DNA (deoxyribonucleic acid) immobilization and hybridization. The negatively charged probe single-stranded DNA (ssDNA) molecules were electrostatically adsorbed onto the positively charged PAH layer, resulting in a preferentially flat orientation of the ssDNA molecules within the Debye length, thus yielding a reduced charge-screening effect and a higher sensor signal. Each sensor-surface modification step (PAH adsorption, probe ssDNA immobilization, hybridization with complementary target DNA (cDNA), reducing an unspecific adsorption by a blocking agent, incubation with noncomplementary DNA (ncDNA) solution) was monitored by means of capacitance–voltage and constant-capacitance measurements. In addition, the surface morphology of the PAH layer was studied by atomic force microscopy and contact-angle measurements. High hybridization signals of 34 and 43 mV were recorded in low-ionic strength solutions of 10 and 1 mM, respectively. In contrast, a small signal of 4 mV was recorded in the case of unspecific adsorption of fully mismatched ncDNA. The density of probe ssDNA and dsDNA molecules as well as the hybridization efficiency was estimated using the experimentally measured DNA immobilization and hybridization signals and a simplified double-layer capacitor model. The results of field-effect experiments were supported by fluorescence measurements, verifying the DNA-immobilization and hybridization event.
Designing novel or optimizing existing biodegradable polymers for biomedical applications requires numerous tests on the effect of substances on the degradation process. In the present work, polymer-modified electrolyte–insulator–semiconductor (PMEIS) sensors have been applied for monitoring an enzymatically catalyzed degradation of polymers for the first time. The thin films of biodegradable polymer poly(d,l-lactic acid) and enzyme lipase were used as a model system. During degradation, the sensors were read-out by means of impedance spectroscopy. In order to interpret the data obtained from impedance measurements, an electrical equivalent circuit model was developed. In addition, morphological investigations of the polymer surface have been performed by means of in situ atomic force microscopy. The sensor signal change, which reflects the progress of degradation, indicates an accelerated degradation in the presence of the enzyme compared to hydrolysis in neutral pH buffer media. The degradation rate increases with increasing enzyme concentration. The obtained results demonstrate the potential of PMEIS sensors as a very promising tool for in situ and real-time monitoring of degradation of polymers.
Label-free sensing of biomolecules by their intrinsic molecular charge using field-effect devices
(2015)
A multi-spot light-addressable potentiometric sensor (LAPS), which belongs to the family of semiconductor field-effect devices, was applied for label-free detection of double-stranded deoxyribonucleic acid (dsDNA) molecules by their intrinsic molecular charge. To reduce the distance between the DNA charge and sensor surface and thus, to enhance the electrostatic coupling between the dsDNA molecules and the LAPS, the negatively charged dsDNA molecules were electrostatically adsorbed onto the gate surface of the LAPS covered with a positively charged weak polyelectrolyte layer of PAH (poly(allylamine hydrochloride)). The surface potential changes in each spot of the LAPS, induced by the layer-by-layer adsorption of a PAH/dsDNA bilayer, were recorded by means of photocurrent-voltage and constant-photocurrent measurements. In addition, the surface morphology of the gate surface before and after consecutive electrostatic adsorption of PAH and dsDNA layers was studied by atomic force microscopy measurements. Moreover, fluorescence microscopy was used to verify the successful adsorption of dsDNA molecules onto the PAH-modified LAPS surface. A high sensor signal of 25 mV was registered after adsorption of 10 nM dsDNA molecules. The lower detection limit is down to 0.1 nM dsDNA. The obtained results demonstrate that the PAH-modified LAPS device provides a convenient and rapid platform for the direct label-free electrical detection of in-solution hybridized dsDNA molecules.
The conjunction of (bio-)chemical recognition elements with nanoscale biological building blocks such as virus particles is considered as a very promising strategy for the creation of biohybrids opening novel opportunities for label-free biosensing. This work presents a new approach for the development of biosensors using tobacco mosaic virus (TMV) nanotubes or coat proteins (CPs) as enzyme nanocarriers. Sensor chips combining an array of Pt electrodes loaded with glucose oxidase (GOD)-modified TMV nanotubes or CP aggregates were used for amperometric detection of glucose as a model system for the first time. The presence of TMV nanotubes or CPs on the sensor surface allows binding of a high amount of precisely positioned enzymes without substantial loss of their activity, and may also ensure accessibility of their active centers for analyte molecules. Specific and efficient immobilization of streptavidin-conjugated GOD ([SA]-GOD) complexes on biotinylated TMV nanotubes or CPs was achieved via bioaffinity binding. These layouts were tested in parallel with glucose sensors with adsorptively immobilized [SA]-GOD, as well as [SA]-GOD crosslinked with glutardialdehyde, and came out to exhibit superior sensor performance. The achieved results underline a great potential of an integration of virus/biomolecule hybrids with electronic transducers for future applications in biosensorics and biochips.