In this work I present a novel approach to the analysis of biomolecules, and a study on two derived practical applications to evaluate its constraints, limits, and potential benefits, namely a biosensing device and a selective transport through membrane.
The new approach is based on a 100-800 nm pore etched in a silicon nitride membrane. A linear target molecule, such as DNA, is inserted in the pore and linked at both termini with anchors, one on each side of the pore. Since the complex is stable and the linked objects have a size that is much larger than the target molecule, manipulation, pore closure/opening, possible interactions, stretching and other forces, and in general several characteristics and behaviours of the molecules can be studied at the pore interface. The realization of such a device is preliminary to the development of novel pore-based analytical tools.
The principle was applied for the development of a biosensing device. Biosensing devices that perform electrical signal detection are facing the need of being both extremely small and highly sensitive, that is particularly challenging for conventional biosensors where the signal produced is proportional to the surface detecting area. Here, I report the production of a sensor device based on DNA specific displacement of a stable blockade in a synthetic pore section, due to objects associated with the interacting molecules. Thus the signal is proportional to the pore size and not to the surface containing the target/probe molecule.
First, I report the setting up of the single components of the device: a complex made of a DNA linker and two particles –the anchors-, the synthetic nanopored membrane and an electrophoretic cell together with an electromagnet -the sensing tools-. Then I show the results of trans-membrane interactions between the objects both outside and inside the sensor device. The applications results related to the biosensor operation are then shown, reporting the detection of the hybridization or the strand-displacement between probes and targets DNA molecules. Finally, I show the operation of a trans-membrane transporter mediated by particles carriers, where the system is exploited to capture and import target molecules through the membrane.
