A study of bioreactor surfaces for cell interactions and sensing.
Abstract
Polymeric hydrogels were used to create bio-smart hydrogels serving multifunctional
roles interfacing with cells and enzyme substrates. Their value lies in their
use as:
i) Stimuli- responsive membranes that directly transmute chemical potential energy
into proportionate electrical signals, ii) as biomimetically inspired biocompatible
coatings on stents and other implantable bionic devices, iii) as bio receptor hosting
membranes for enzyme-based implantable biosensors. Biosensors use oxidoreductase
enzymes such as glucose oxidase (GOx) and lactate oxidase (LOx) to confer
specificity. Such enzymes may initiate more complex in vivo inflammatory response.
In this thesis individual and combined effects of different enzymes (GOx,
Superoxide dismutase (SOD), and catalase) were studied to achieve hydrogelenzyme
systems, which in theory may mitigate against adverse cell outcomes.
The incorporation of enzymes into bioactive hydrogels was investigated, and
revealed effects on the growth, viability and attachment of surface dependant
RMS13 human muscle fibroblasts and B50 rat neuronal cells. Agarose and
p(HEMA)-based hydrogels were prepared with fibrinogen 5% (w/v) to promote
integrin-mediated cellular attachment and also with different combinations of
glucose oxidase (GOx), catalase (CAT) and superoxide dismutase (SOD). Cell
viability was maintained best on catalase hydrogels. The presence of GOx within
hydrogels membrane compromised cell viability in both hydrogel types, presumably
due to accumulation of H2O2 confirmed by amperometric detection using fabricated
platinum needle electrodes. Hydrogels prepared with GOx and CAT showed improved cell viability, further suggesting the negative influence of H2O2. High
temperature treatment of the enzyme-hydrogel membranes, resulting in enzyme
denaturation, returned all constructs to control levels of viability, confirming the
relationship of cell viability with enzyme activity.
An additional study was undertaken into the viability and growth of B50 cells on
crosslinked protein membranes of fibrinogen and albumin as a potential bioreactor
surface. The use of crosslinked fibrinogen to facilitate cell growth within
microfluidic channels appears to have been realized. Fabrication and use of
miniaturized gold-filled silica recess and inlaid disc electrodes, compared with the
use of agarose gels in the recesses was investigated to improve stabilization of an
amperometric H2O2 electrode. From this, a microfluidic device with an integrated
inner diameter working and counter / reference electrode was fabricated which
showed feasibility of more rapid amperometric detection of H2O2 in miniature flow
channels.
Authors
Khan, Rachel MarinaCollections
- Theses [3822]