The filamentous peptide-based nanowires produced by some dissimilatory metal-reducing bacteria, such as Geobacter sulfurreducens, display excellent natural conductivity. Their mechanism of conduction is assumed to be a combination of delocalized electrons through closely aligned aromatic amino acids and hopping/charge transfer. The proteins that form these microbial nanowires are structured from a coiled-coil, for which the design rules have been reported in the literature. Furthermore, at least one biomimetic system using related synthetic peptides has shown that the incorporation of aromatic residues can be used to enhance conductivity of peptide fibers. Herein, the de novo design of peptide sequences is used to enhance the conductivity of peptide gels, as inspired by microbial nanowires. A critical factor hampering investigations in both microbiology and materials development is inconsistent reporting of biomaterial conductivity measurements, with consistent methodologies needed for such investigations. We have reported a method herein to analyze non-Ohmic behavior using existing parameters, which is a statistically insightful approach for detecting small changes in biologically based samples. Aromatic residues were found to contribute to peptide gel conductivity, with the importance of the peptide confirmation and fibril assembly demonstrated both experimentally and computationally. This is a small step (in combination with parallel research under way by other researchers) toward developing effective peptide-based conducting nanowires, opening the door to the use of electronics in water and physiological environments for bioelectronic and bioenergy applications.
Transfection cell microarrays (TCMs) are a high-throughput, miniaturised cell-culture system utilising reverse transfection, in which cells are seeded onto a DNA array resulting in localised regions of transfected cells. TCMs are useful for the analysis of gene expression, and can be used to identify genes involved in many cellular processes. This is of significant interest in fields such as tissue engineering, diagnostic screening, and drug testing[1, 2]. Low transfection efficiency has so far limited the application and utility of this technique. Recently, the transfection efficiency of TCMs was improved by an application of a high voltage for a short period of time to the DNA array resulting in the electroporation of cells attached to the surface[3, 4]. Furthermore, application of a low voltage for a longer period of time to the DNA array was shown to improve the transfection efficiency by stimulating the desorption of attached DNA, increasing the concentration of DNA available for cellular uptake[5]. In the present study, the optimisation of the uptake of adsorbed DNA vectors by adherent cells, utilising a voltage bias without compromising cell viability was investigated. This was achieved by depositing negatively charged DNA plasmids onto a positively charged allylamine plasma polymer (ALAPP) layer deposited on highly doped p-type silicon wafers either using a pipettor or a microarray contact printer. Surface-dependant human embryonic kidney (HEK 293 line) cells were cultured onto the DNA vector loaded ALAPP spots and the plasmid transfection events were detected by fluorescence microscopy. Cell viability assays, including fluorescein diacetate (FDA) / Hoechst DNA labelling, were carried out to determine the number of live adherent cells before and after application of a voltage. A protocol was developed to screen for voltage biases and exposure times in order to optimise transfection efficiency and cell viability. Cross-contamination between the microarray spots carrying different DNA vectors was also investigated. By application of a voltage of 286 V/cm for 10 ms, transfection efficiency was doubled compared to using only transfection reagent, whilst maintaining a cell viability of 60-70% of the positive control.
A new class of peptide-based dendrons and dendrimers is presented that display unique vesicle-driven organogelation. We demonstrate the crosslinking of diacetylene-based dendrimers in the gel state, producing strongly fluorescent polydiacetylenes. The crosslinked gels crafted from peptide dendrimers may find practical use in various applications, including as scaffold materials in tissue engineering and for drug delivery.
The spatial control over biomolecule- and cell-surface interactions is of great interest to a broad range of biomedical applications, including sensors, implantable devices and cell microarrays. Microarrays in particular require precise spatial control and the formation of patterns with microscale features. Here, we have developed an approach specifically designed for transfected cell microarray (TCM) applications that allows microscale spatial control over the location of both DNA and cells on highly doped p-type silicon substrates. This was achieved by surface modification, involving plasma polymerization of allylamine, grafting of poly(ethylene glycol) and subsequent excimer laser ablation. DNA could be delivered in a spatially defined manner using ink-jet printing. In addition, electroporation was investigated as an approach to transfect attached cells with adsorbed DNA and good transfection efficiencies of approximately 20% were observed. The ability of the microstructured surfaces to spatially direct both DNA adsorption and cell attachment was demonstrated in a functional TCM, making this system an exciting platform for chip-based functional genomics.
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