Bioelectronic Circuits Research Articles

Temperature Dependence of Electrochemical DNA Charge Transport: Influence of a Mismatch

Charge transfer through DNA is of interest as DNA is both the quintessential biomolecule of all living organisms and a self-organizing element in bioelectronic circuits and sensing applications. Here, we report the temperature-dependent properties of DNA charge transport in an electronically relevant arrangement of DNA monolayers on gold under biologically relevant conditions, and we track the effects of incorporating a CA single base pair mismatch. Charge transfer (CT) through double stranded, 17mer monolayers was monitored by following the yield of electrochemical reduction of a Nile blue redox probe conjugated to a modified thymine. Analysis with cyclic voltammetry and square wave voltammetry shows that DNA CT increases significantly with temperature, indicative of more DNA bridges becoming active for transport. The mismatch was found to attenuate DNA CT at lower temperatures, but the effect of the mismatch diminished as temperature was increased. Voltammograms were analyzed to extract the electron transfer rate k(0), the electron transfer coefficient α, and the redox-active surface coverage Γ*. Arrhenius behavior was observed, with activation energies of 100 meV for electron transfer through well-matched DNA. Single CA mismatches increased the activation energy by 60 meV. These results have clear implications for sensing applications and are evaluated with respect to the prominent models of DNA CT.

Single molecule recordings of lysozyme activity.

Single molecule bioelectronic circuits provide an opportunity to study chemical kinetics and kinetic variability with bond-by-bond resolution. To demonstrate this approach, we examined the catalytic activity of T4 lysozyme processing peptidoglycan substrates. Monitoring a single lysozyme molecule through changes in a circuit's conductance helped elucidate unexplored and previously invisible aspects of lysozyme's catalytic mechanism and demonstrated lysozyme to be a processive enzyme governed by 9 independent time constants. The variation of each time constant with pH or substrate crosslinking provided different insights into catalytic activity and dynamic disorder. Overall, ten lysozyme variants were synthesized and tested in single molecule circuits to dissect the transduction of chemical activity into electronic signals. Measurements show that a single amino acid with the appropriate properties is sufficient for good signal generation, proving that the single molecule circuit technique can be easily extended to other proteins.

A protein transistor made of an antibody molecule and two gold nanoparticles

A major challenge in molecular electronics is to attach electrodes to single molecules in a reproducible manner to make molecular junctions that can be operated as transistors. Several attempts have been made to attach electrodes to proteins, but these devices have been unstable. Here, we show that self-assembly can be used to fabricate, in a highly reproducible manner, molecular junctions in which an antibody molecule (immunoglobulin G) binds to two gold nanoparticles, which in turn are connected to source and drain electrodes. We also demonstrate effective gating of the devices with an applied voltage, and show that the charge transport characteristics of these protein transistors are caused by conformational changes in the antibody. Moreover, by attaching CdSe quantum dots to the antibody, we show that the protein transistor can also be gated by an applied optical field. This approach offers a versatile platform for investigations of single-molecule-based biological functions and might also lead to the large-scale manufacture of integrated bioelectronic circuits.

Self-assembled dendritic nanowires of Au–Pt alloy through electrodeposition from solution under AC fields

Au–Pt dentritic nanowires (DNWs) bridging a pair of 10–40μm spacing electrodes were successfully self-assembled from the corresponding ions solution via deposition of an alternating current (AC). The morphology and the composition of Au–Pt DNWs were characterized by scanning electron microscopy (SEM) and energy-dispersion X-ray (EDX) spectrometer. In the electrolyte of various molar ratios of [AuCl4]−/[PtCl6]2−, Au–Pt DNWs with 8–95at% Pt were fabricated through applying a constant voltage 8–16Vpp and varied frequencies of 300Hz for nucleation and 1MHz for growth. The DNWs have the diameter range of 100–200nm. The X-ray diffraction (XRD) results confirmed that the DNWs of 20 or 80at% Pt have the alloy phase structure. Au–Pt DNWs are promising nanomaterials of chemical sensors, bioelectronic circuits and nanoelectronic interconnection.

Biomass Champo leaf transistor configuration

Dynamics of charged nano particles of green leaf sap or photo synthesised liquid food under interaction of external EMF plays a vital role in the functioning of biomass electrical circuits. Using this concept, we for the first time realised a biomass Champo leaf transistor configuration. The functioning of bio electronic circuit components resistance R, capacitance C and diode have been earlier reported. From the theory of electronics, a transistor consist of two electronic diode connected back to back. This concept was utilised to realise biomass Champo leaf transistor configuration. The measured input characteristics (V BE vs I BE ) and output characteristics (V CE vs. I CE ) along with β are given. Applications towards plant/tree health, soil moisture monitoring and strategic communication are envisaged.

Processing and quantitative analysis of biodegradable polymers (PLLA and PCL) thermal bonding

A quantitative analysis of the bond strength and microstructure integrity achieved when bonding the biodegradable polymers poly(L-lactide) (PLLA) and poly(ε-caprolactone) (PCL) has been performed using the response surface methodology. The respective influence of the bonding parameters (temperature, pressure, duration) on the bond strength and microchannel integrity was investigated. PLLA and PCL were identified as suitable candidates for packaging materials for bioelectronic circuits of conductive biodegradable polymers. For a future packaging application, the bonding parameters were adapted to optimize the bond strength; the estimated values for the bond strength and channel integrity that were predicted by the surface plots were 2.32 ± 0.26 MPa and 33.7 ± 12.9% for PLLA, and 0.81 ± 0.11 MPa and 50.9 ± 5.7% for PCL. These values were in good agreement with the experimentally determined bond strength of 2.00 ± 1.10 MPa (PLLA) and 0.67 ± 0.22 MPa (PCL) and deformation of 31.4 ± 7.0% (PLLA) and 52.9 ± 4.1% (PCL). Microchannels with an aspect ratio of 1:12.5 were successfully fabricated. The impact of the fabrication process on the PLLA and PCL chemical properties was also investigated through differential scanning calorimetry and gel permeation chromatography measurements. It was observed that the weight average molecular weight Mw decreases after each fabrication step, as much as 68% for PLLA and 59% for PCL. The strongest reduction was observed after the compression molding (above the melting temperature) which should be kept as short as possible. An annealing step allowed increasing the crystallinity and improved the overall polymer stiffness.

Ion bipolar junction transistors

Dynamic control of chemical microenvironments is essential for continued development in numerous fields of life sciences. Such control could be achieved with active chemical circuits for delivery of ions and biomolecules. As the basis for such circuitry, we report a solid-state ion bipolar junction transistor (IBJT) based on conducting polymers and thin films of anion- and cation-selective membranes. The IBJT is the ionic analogue to the conventional semiconductor BJT and is manufactured using standard microfabrication techniques. Transistor characteristics along with a model describing the principle of operation, in which an anionic base current amplifies a cationic collector current, are presented. By employing the IBJT as a bioelectronic circuit element for delivery of the neurotransmitter acetylcholine, its efficacy in modulating neuronal cell signaling is demonstrated.

Characterization Methods of a Whole-Cell Bioluminescent Biosensor

A whole-cell bioluminescent biosensor dynamics based on genetically engineered Escherichia coli bacteria carrying a promoter-reporter fusion is often determined by a series of chemical reactions such as gene expression and reactions involving protein concentrations at nanomolar volume. In this paper, we derive an anayltical model of a whole-cell bioluminescent biosensor based on the Michaelis-Menten kinetics of simple enzymatic reactions. The proposed model is characterized by three measurable set of parameters: the biosensor effective rate constant, the total number of the emitted photons and the biosensor reaction order. We get a good agreement between the simulation results of the model and the measured light signal, correlating the input signal (toxin concentration) and the output signal (the bioluminescent light). The model is tested and verified for various prompters (recA, KatG, and micF). This model is useful in the design process of bio-electronic circuits.

Synthesis and room temperature photo-induced electron transfer in biologically active bis(terpyridine)ruthenium(ii)–cytochrome c bioconjugates and the effect of solvents on the bioconjugation of cytochrome c

Photo-active bis(terpyridine)ruthenium(ii) chromophores were synthesised and attached to the redox enzyme iso-1 cytochrome c in a mixed solvent system to form photo-induced bioconjugates in greater than 40% yield after purification. The effects of up to 20% (v/v) of acetonitrile, tetrahydrofuran, dimethylformamide, or dimethyl sulfoxide at 4, 25 and 35 degrees C on the stability and biological activity of cytochrome c and its reactivity towards the model compound 4,4'-dithiodipyridine (DTDP) was measured. The second-order rate constant for the DTDP reaction was found to range between k = 2.5-4.3 M(-1) s(-1) for reactions with 5% organic solvent added compared to k = 5.6 M(-1) s(-1) in pure water at 25 degrees C. Use of 20% solvent generally results in significant protein oxidation, and 20% acetonitrile and tetrahydrofuran in particular result in significant protein dimerization, which competes with the bioconjugation reaction. Cyclic voltammetry studies indicated that the rate of electron transfer to the heme in solution was reduced in the bis(terpyridine)ruthenium(ii) cytochrome c bioconjugates compared to unmodified cytochrome c. Steady-state fluorescence studies on these bioconjugates showed that energy or electron transfer is taking place between the bis(terpyridine)ruthenium(ii) chromophores and cytochrome c. The bis(terpyridine)ruthenium(ii) cytochrome c bioconjugates demonstrate room temperature photo-activated electron transfer from the bis(terpyridine)ruthenium(ii) donor to the protein acceptor. Two sacrificial donors were used; in 50% glycerol, the bioconjugates were reduced in about 15 min while in 20 mM EDTA the bioconjugates were fully reduced in less than 5 min upon irradiation with a xenon lamp source. Under these conditions, the reduction of the non-covalent mixture of cytochrome c and bis(terpyridine)ruthenium(ii) mixtures took over 30 min. Control experiments showed that the photo-induced reduction of cytochrome c only occurs in the absence of oxygen and presence of a sacrificial donor. These results are encouraging for future incorporation of these bioconjugates in light-responsive bioelectronic circuits, including photo-activated biosensors and biofuel cells.

Evaluation of the Photosynthetic Reaction Center Protein for Potential Use as a Bioelectronic Circuit Element

The characterization of a bioelectronic composite prepared by molecular wiring of a bacterial photosynthetic reaction center (RC) to a metal (Au) electrode is described. Two unique attachment sites on the protein surface were studied as sites for electrical connections--a polyhistidine tag introduced by site-directed mutagenesis and a native cysteine amino acid residue. These two attachment sites were evaluated independently and found to serve effectively in coupling the protein to the electrode surface asymmetrically. Cyclic voltammetry (CV) was used to monitor protein integrity and confirm protein chemisorption and orientation to the organofunctionalized gold electrode. Single-protein transport measurements made with conductive atomic force microscopy (C-AFM) were used to study the electrical transport. Current-voltage (I-V) curves obtained by wiring the protein at the polyhistidine tag showed diodelike behavior. The cysteine attachment site does not serve as an efficient means to address the protein electrically. Scanning tunneling spectroscopy (STS) performed on RCs coupled at the donor side under both dark- and white-light-illuminated conditions confirmed the C-AFM studies.

Evaluation of the Photosynthetic Reaction Center Protein for Potential Use as a Bioelectronic Circuit Element

Evaluation of the Photosynthetic Reaction Center Protein for Potential Use as a Bioelectronic Circuit Element

Microcontact printing of metalloproteins

The morphological investigation by scanning probes techniques, and especially Atomic Force Microscopy (AFM) allow to measure the surface roughness of immobilized proteins with unequalled vertical spatial resolution. This is particularly important, since the biological role of proteins is closely related to their physiological folding architectures. In particular, metalloproteins, such as blue copper proteins, revealed to be good candidates for biomolecular electronics by virtue of their natural electron transfer activity, which is exploitable in molecular switches whose conduction state can be controlled by gating their redox state. The interest in these molecules is indeed motivated by the possibility of eliciting current flow through the redox level of single molecules. Printing metalloproteins can be an advantageous approach for the easy realization of bioelectronic circuits. We implemented and developed the selective deposition of the metalloprotein Azurin (Az) from Pseudomonas aeruginosa by microcontact printing, and investigated by AFM the surface morphology of the printed Az layers.

Bonding silicon-on-insulator to glass wafers for integrated bio-electronic circuits

We report a method for bonding silicon-on-insulator wafers onto glass wafers. After pre-cleaning the wafers by an ozone and ultraviolet exposure, followed by mega-sonic water rinse, the SOI wafers are bonded to glass wafers in a vacuum chamber. This is performed at a temperature of 400 °C under an applied voltage of 700 V. The interface between the glass and SOI wafer is tested mechanically and inspected by electron beam microscopy. Furthermore, we demonstrate removal of the silicon bulk layer after wafer bonding. The quality of the single crystalline Si thin film on the glass wafers has been verified by four-circle x-ray diffraction and scanning electron microscopy. This process will allow us the integration of thin-film electronics in biological sensor applications.

Dielectrophoretic assembly of electrically functional microwires from nanoparticle suspensions.

A new class of microwires can be assembled by dielectrophoresis from suspensions of metallic nanoparticles. The wires are formed in the gaps between planar electrodes and can grow faster than 50 micrometers per second to lengths exceeding 5 millimeters. They have good ohmic conductance and automatically form electrical connections to conductive islands or particles. The thickness and the fractal dimension of the wires can be controlled, and composite wires with a metallic core surrounded by a latex shell can be assembled. The simple assembly process and their high surface-to-volume ratio make these structures promising for wet electronic and bioelectronic circuits.