Simulteanous enzyme immobilisation and micro-nanofabrication process for low-cost biosensor applications

Biosensors which are based on BioMEMS combine developments in micro-nanotechnology and biological recognition elements, in order to enable the measurement of low level analyte concentrations with portable detection-systems. A major challenge of such small biochips is the accuracy of the sensing process. The main aim of the present research project was the investigation and development of novel low cost matrices based on custom-made SU-8 polymer for simultaneous microfabrication and immobilisation of enzymes in order to form the basis for the development of novel low-cost micro-nano-biosensors. The negative photoresist SU-8 was utilised because of its attractive characterestics such as stability under environmental conditions, biocompatibility and low cost. The entrapment of biological molecules into the SU-8 structures was experimentally assessed using the enzyme glucose oxidase; and, electrochemical and optical sensing techniques were employed. The major emphasis of the experimental studies was on selectivity, sensitivity and stability improvements as well as on the optimum design, and microfabrication process. A detailed investigation of the optimum microfabrication process and composition of biocompatible SU-8 matrices with encapsulated enzyme such as glucose oxidase was carried out. The conditions of the various steps of the SU-8 processing are hostile to the enzymes’ survival. Improving the process conditions and encapsulation matrices were primordial for the success of this work. This study implied variation in enzyme concentration matrices in order to improve the activity of the encapsulated enzyme inside the matrix. Different sacrificial materials for SU-8 and their influence on the used enzyme and on the products of enzymatic reactions were investigated.

PROOF OF CONCEPT OF A NOVEL OPTICAL GLUCOSE ENZYME-BASED MICRO-BIOSENSOR USING SU-8

Nico de Rooij, Sotiria Psoma, Peter Van der Wal

A novel one-step microfabrication process of SU-8 films for the development of inexpensive light-weight and small size optical glucose micro-biosensors is proposed. Experimental work was carried out in order to investigate whether SU-8 photoresists, which are polymers widely used in MEMS applications, can be utilised as immobilisation matrices for the simultaneous encapsulation of glucose oxidase and an oxygen-sensitive fluorescent indicator. The immobilisation of the enzyme, the encapsulation of the indicator and the patterning of the custom-made SU-8 take place simultaneously, thus simplifying the microfabrication process.

2011

A novel enzyme entrapment in SU-8 microfabricated films for glucose micro-biosensors

Olivier Frey, Sotiria Psoma

The present work investigates the utilisation of the widely used SU-8 photoresist as an immobilisation matrix for glucose oxidase (GOx) for the development of glucose micro-biosensors. The strong advantage of the proposed approach is the simultaneous enzyme entrapment during the microfabrication process within a single step, which is of high importance for the simplification of the BioMEMS procedures. Successful encapsulation of the enzyme GOx in "customised" SU-8 microfabricated structures was achieved through optimisation of the one-step microfabrication process. Although the process involved contact with organic solvents, UV-light exposure, heating for pre- and post-bake and enzyme entrapment in a hard and rigid epoxy resin matrix, the enzyme retained its activity after encapsulation in SU-8. Measurements of the immobilised enzyme's activity inside the SU-8 matrix were carried out using amperometric detection of hydrogen peroxide in a 3-electrode setup. Films without enzyme showed negligible variation in current upon the addition of glucose, as opposed to films with encapsulated enzyme which showed a very clear increase in current. Experiments using films of increased thickness or enzyme concentration, showed a higher response, thus proving that the enzyme remained active not only on the film's surface, but also inside the matrix as well. The proposed enzyme immobilisation in SU-8 films opens up new possibilities for combining BioMEMS with biosensors and organic electronics. (C) 2010 Elsevier B.V. All rights reserved.

Elsevier2010

Fluorescence based low-cost microbiosensors using simultaneously microfabrication process and enzyme immobilisation

Nico de Rooij, Sotiria Psoma, Peter Van der Wal

A novel one-step microfabrication process of SU-8 films is proposed for developing inexpensive, small size and light-weight optical glucose micro-biosensors. Experimental work was performed in order to investigate whether the widely used in MEMS applications SU-8 photoresists polymers, can be utilised as immobilisation matrices for the simultaneous encapsulation of an oxygen-sensitive fluorescent indicator and glucose oxidase. The enzyme immobilisation, the encapsulation of the indicator and the patterning of the SU-8 take place simultaneously, thus offering a significant simplification of the microfabrication process. Despite the process involved contact with organic solvents, UV-light exposure and heating for pre- and post-bake, and the embedding of the enzyme in a hard and rigid epoxy resin matrix, it was observed that the enzyme demonstrated activity after encapsulation in SU-8. Testing of the immobilised enzyme´s activity inside the SU-8 matrix, was carried out with the measurement of oxygen consumption using an oxygen-sensitive indicator during the enzymatic oxidation of glucose. Negligible variation in fluorescence intensity upon the addition of glucose was observed in films without enzyme, whereas films with encapsulated enzyme and oxygen-sensitive fluorescent indicator showed a very clear increase in fluorescence intensity upon addition of glucose. A high transparency and negligible fluorescence of the SU-8 films were observed. The presented work opens up new possibilities for combining BioMEMS, advanced optoelectronic components with smart biosensor technology for personalised medicine diagnostic applications.

2012