In recent years, microRNA (miRNA) has garnered a high level of interest in the field of
biosensor development. MiRNA are a class of small, circulating RNA sequences that
are essential for healthy control of protein expression. The variation of levels of specific
miRNAs has been linked with over 150 diseases since the turn of the millennium,
including cancers, cardiovascular diseases, parasitic infections and neurological
disorders. The improved prognosis from early detection is stark, but established
methods of miRNA detection suffer from poor sensitivity, low throughput, and require
specialised laboratory equipment and trained staff to perform the time-consuming
techniques. A simple, cheap and sensitive miRNA point of care sensor would be an
invaluable tool in healthcare.
This thesis presents the continued optimisation of a miRNA sensing hydrogel with
oligonucleotide crosslinks that are selectively cleaved in the presence of the target
miRNA sequence. This selective reduction in crosslink density was transduced by a
change in the swelling profile of the hydrogel and intelligent crosslink design used to
control the swelling response for detecting a miRNA sequence, a short RNA (sRNA)
sequence, or a small molecule using an aptamer. Morpholino oligonucleotides (MOs),
an uncharged DNA analogue, were functionalised with an acrylamide moiety and used
as responsive crosslinks for miRNA sequence detection in a world’s first MO
crosslinked hydrogel. The MO crosslinks offered significant improvements over DNA
crosslinked hydrogels through improved thermal stability, no salt requirement and
1000-fold improved sensitivity, facilitating a wider range of sensing conditions.
Analysis was also achieved using a mobile phone camera and laptop, demonstrating
portability.
Carbon nanoparticles (CNP) were suspended in the hydrogels to act as a conductive
component. As the hydrogel swells the distance between the particles is increased
until there is no conductive pathway, resulting in an increase in the hydrogel’s
resistance. Numerous reproducibility challenges were identified with regards to gel
delamination and CNP leaching partly due to inefficient UV photoinitiation of the
pigment composite pre-gel solution. SEM imaging identified inconsistent composite
homogeneity with areas of higher CNP and gel density in DNA crosslinked
composites, while MO crosslinked composites were homogenous and less
conductive.
Inkjet printing of the composite material using an electrostatic dispersion as the
conductive component was made possible using MO crosslinks with no salt and
ammonium persulfate with TEMED in place of UV initiation. Optimised synthesis
resulted in homogenous conductive composites far more robust and reproducible than
the UV initiated CNP composite. However, MO solution viscosity resulted in improper
aspiration and inaccurate deposition. Potential solutions and improvements are
suggested, facilitated by the improvements offered by MO crosslinks
