RNA interference (RNAi) methods for insects are often limited by problems with double-stranded (ds) RNA delivery, which restricts reverse genetics studies and the development of RNAi-based biocides. We therefore delegated to insect symbiotic bacteria the task of: (i) constitutive dsRNA synthesis and (ii) trauma-free delivery. RNaseIII-deficient, dsRNA-expressing bacterial strains were created from the symbionts of two very diverse pest species: a long-lived blood-sucking bug, Rhodnius prolixus, and a short-lived globally invasive polyphagous agricultural pest, western flower thrips (Frankliniella occidentalis). When ingested, the manipulated bacteria colonized the insects, successfully competed with the wild-type microflora, and sustainably mediated systemic knockdown phenotypes that were horizontally transmissible. This represents a significant advance in the ability to deliver RNAi, potentially to a large range of non-model insects.
A hybrid biosensor based on a graphene resistor functionalized with self-assembled Graphene-AuNPs (Gold Nanoparticles) is demonstrated for the real-time detection of hepatitis B surface antigen (HBsAg). The hybrid biosensor consists of a ssDNA sequence attached to a graphene resistor device via π–π stacking interactions in combination with a ssDNA functionalized AuNP. The ssDNA has complementary sequences which through hybridization, yield the graphene-AuNP hybrid biosensor. Real-time 2-point resistance measurements, performed using varying concentrations of HBsAg, show a linear dependence of resistance change against the logarithm of HBsAg concentration (log[HBsAg]). A limit of detection of 50 pg ml−1 was observed. Moreover, the hybrid biosensor platform has potential to be applied to any biomarker of interest.
Affinity biosensors based on graphene field-effect transistor (GFET) or resistor designs require the utilization of graphene’s exceptional electrical properties. Therefore, it is critical when designing these sensors, that the electrical properties of graphene are maintained throughout the functionalization process. To that end, non-covalent functionalization may be preferred over covalent modification. Drop-cast 1,5-diaminonaphthalene (DAN) was investigated as a quick and simple method for the non-covalent amine functionalization of carbon-based surfaces such as graphene, for use in biosensor development. In this work, multiple graphene surfaces were functionalized with DAN via a drop-cast method, leading to amine moieties, available for subsequent attachment to receptor molecules. Successful modification of graphene with DAN via a drop-cast method was confirmed using X-ray photoelectron spectroscopy (XPS), Raman spectroscopy and real-time resistance measurements. Successful attachment of receptor molecules also confirmed using the aforementioned techniques. Furthermore, an investigation into the effect of sequential wash steps which are required in biosensor manufacture, on the presence of the DAN layer, confirmed that the functional layer was not removed, even after multiple solvent exposures. Drop-cast DAN is thus, a viable fast and robust method for the amine functionalization of graphene surfaces for use in biosensor development.
Graphene-based point-of-care (PoC) and chemical sensors can be fabricated using photolithographic processes at wafer-scale. However, these approaches are known to leave polymer residues on the graphene surface, which are difficult to remove completely. In addition, graphene growth and transfer processes can introduce defects into the graphene layer. Both defects and resist contamination can affect the homogeneity of graphene-based PoC sensors, leading to inconsistent device performance and unreliable sensing. Sensor reliability is also affected by the harsh chemical environments used for chemical functionalisation of graphene PoC sensors, which can degrade parts of the sensor device. Therefore, a reliable, wafer-scale method of passivation, which isolates the graphene from the rest of the device, protecting the less robust device features from any aggressive chemicals, must be devised. This work covers the application of molecular vapour deposition technology to create a dielectric passivation film that protects graphene-based biosensing devices from harsh chemicals. We utilise a previously reported “healing effect” of Al2O3 on graphene to reduce photoresist residue from the graphene surface and reduce the prevalence of graphene defects to improve graphene device homogeneity. The improvement in device consistency allows for more reliable, homogeneous graphene devices, that can be fabricated at wafer-scale for sensing and biosensing applications.
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