The ability to utilise new knowledge of biomarkers from genomic and proteomic data
will have a great impact on molecular diagnosis. Biomarker detection could be
achieved by utilising a capture molecule that associates specifically with the target
biomarker. The work described in this thesis focuses on a platform comprising a
lysozyme binding aptamer and an amperometric electrode (an electrochemical
aptasensor). To couple the binding reaction to a change in current, the aptamer is
modified with a redox group, ferrocene. Two types of signalling aptamer were
constructed, one comprised the aptamer self-assembled on gold and hybridised to a
short complementary oligonucleotide carrying a ferrocene group. The second
incorporated the binding sequence into a molecular beacon, one end of which self-assembled
onto the electrode, the other end carried the ferrocene group. Both of these
showed a lysozyme dependent change in current on a gold electrode.
Further characterisation of the first aptasensor suggested that the nucleic acid formed
a multilayer structure on the electrode surface and that lysozyme binding induced
conformational change moved ferrocene close to the surface, increasing the current. In
contrast, the second aptamer usually showed a decrease in current in the presence of
lysozyme suggesting that the binding resulted in the ferrocene moving away from the
surface.
In order to evaluate the possible use of these aptasensors for continuous in vivo
measurement, needle shaped microelectrodes arrays were produced and the beacon
aptamer immobilised on the surface. These electrodes had high impedance which
resulted in low sensitivity, however lysozyme binding could still be detected using
electrochemical impedance spectroscopy with ferrocyanide in solution. These
microspike arrays could also be used for glucose sensing following modification with
glucose oxidase
