Abstract.

Over the last century, new technologies and techniques have advanced so much that they now affect almost every area of our everyday lives. This is particularly true of the innovations in healthcare, where engineering professionals have been closely involved in medical ventures. From this point of view, the development of new technologies for biomedical microsystems is not only a technological step, but is hardly correlated with many fields of our personal and social life.

Biomedical microsystems include a large class of diagnostic systems for blood or other body fluids analyses and drug delivery systems capable of delivering precise quantities of a drug at the right time and as close to the treatment site as possible. As such, the microfabricated devices utilizing microfluidic subsystems are expected to provide next generation of inexpensive tools for diagnostics. 

Recently, the explosive progress in the fields of cell metabolism and molecular biology has resulted in an increased demand for development of innovative technologies that allow identification and detailed structural studies of bio-molecules to be automatically performed at high speeds and with a high sensitivity. As an example the study of the protein modifications, and resulting repercussions in biological systemspresents unquestionable advantage when performed at the protein level. The traditional methodologies often includes long incubations and extensive manual steps and integrated analysis platforms present large benefits in protein fast identification. Other interesting examples are in utilisation of enzymatic sensors for the food analysis.

Miniaturisation of analytical systems is generally considered to be the strategy that will overcome the requirements of process speed for performing efficient evaluation studies. By utilising the versatility of Silicon micromachining to fabricate efficient minute volume microstructures, it is possible to make analysis systems that are extremely small. The benefits of miniaturisation stem from the increased reaction kinetics in low volumes and the possibility to perform sample-handling procedures at a high speed in micro-litre systems. 

In this contest,the project main goal consists in providing new powerful micro-analytical microsystems based on microfabricated flow-through dispenser for on line micro-litres sample handling and integrated with functionalised microsensors.

For applications in biomedicine and biotechnology of the biomedical microsystems, engineers need to acquire multidisciplinary knowledge. The necessary competencies including BioMEMS microtransducer arrays design and fabrication; microfluidics, sensor surface functionalisation; neural networks, interfaces electronics and system automation.

The present multidisciplinar project is highly innovative, highly challenging and along the lines identified by the European Commission in the framework of the Future EmergingTechnologies Programme. 

Partners:

 

Address	Contact persons
ITC-irst Via Sommarive 18, 38050 Povo (TN)	Dr. M. Zen Dr. L. Lorenzelli Dr. B. Angelini
MiTech Labs - Scuola Superiore Sant'Anna via Carducci 40 - 56127 -Pisa (I)	Prof. P. Dario
Dipartimento di Scienze Chimiche Università di Catania Viale A.Doria 6 - 95125 Catania - Italy	Prof. G. Marletta Prof. A. LicciardelloProf.A. Grassi
CNR-ITB Via Fratelli Cervi 93 20090 Segrate Italy	Dr. Gianluca De Bellis
Tecnologie Innovative di Prodotto Centro Ricerche Fiat Strada Torino 50, 10043 Orbassano (TO)	Dott. Piero Perlo Dr. Maria Margherita Dugand
Biotecnologie e Agricoltura C.R. Casaccia Via Anguillarese 301 00060 S.Maria di Galeria Roma	Dr.Roberto Pilloton

Preliminary Remarks

Over the last decade, new technologies and techniques have advanced so much that they now affect almost every area of our everyday life. This is particularly true of the innovations in healthcare, where engineering professionals have been closely involved in medical ventures. From this point of view, the development of new technologies for biomedical microsystems is not only a technological step, but is hardly correlated with many fields of our personal and social life.

Biomedical microsystems include a large class of diagnostic systems for blood or other body fluids analyses and drug delivery systems capable of delivering precise quantities of a drug at the right time and as close to the treatment site as possible. As such, the microfabricated devices utilising microfluidic subsystems are expected to provide next generation of inexpensive tools for diagnostics. 

Recently, the explosive progress in the fields of cell metabolism and molecular biology has resulted in an increased demand for development of innovative technologies that allow identification and detailed structural study of bio-molecules to be automatically performed at high speeds and with a high sensitivity. As an example, the study of the protein modifications and resulting repercussions in biological systemspresents unquestionable advantage when performed at the molecular level. The traditional methodologies often includes long incubations and extensive manual steps and in this case integrated analysis platforms present large benefits in protein fast identification. Other interesting examples are in utilisation of enzymatic sensors for the food analysis.

Biosensors and bioelectronics represent different points of view of the same technology which is intended for mass production of hybrid devices based on the so called “smart properties” of natural molecules andtechnological materials. Several molecules were extensively studied in the past as functional and active interfaces for sensing or bioelectronic purposes. The most common example is represented by biosensors which are obtained by coupling a biomediator with a transducer. Many natural molecules were purified and used for obtaining both enzyme sensors or immunosensors, anda large spectrum of other natural biomolecules were also investigated, including olfactory receptors and oligonucleotides as sensingelements. Biosensors, biological transduction and biolectronic information storage are the main interesting research areas which will be commercially exploited in the near future. At the moment several biomolecules are used for commercially available analytical devices, but the critical factors for their use are mainly related to their stability and optimal immobilisation, without loss or functional properties, on electronic or optical components. Hybrid, synthetic, natural molecules, including their active fragments or modified derivatives, can be used. Lately, genetic engineered biomolecules seem to be a new and powerful approach for obtaining simpler artificial structures with intact or improved properties, or with additional functional group and activities.Not only biosensing will take in advances for the availability of powerful artificial molecular structures, but also a new generation of microelectronic devices will be certainly affected by this new approach. 

Indeed, miniaturisation of analytical systems is generally considered to be the strategy that will overcome the requirements of process speed for performing efficient evaluation studies. By utilising the versatility of silicon micromachining to fabricate efficient minute volume microstructures, it is possible to make analysis systems that are extremely small. The benefits of miniaturisation stem from the increased reaction kinetics in low volumes and the possibility to perform sample-handling procedures at a high speed in micro-litre systems. 

Microsystems for clinical diagnostics are usually referred to a combination of microchip-sized devices connected together or integrated on a single substrate, (e.g. a chip based silicon biosensor), used for the detection of nucleic acidsor another type of analyte.Reducing cost is also a driving force in molecular diagnostics due to the lack of extensive automation and system integration in clinical chemistry laboratories.

In the effort to create a portable and inexpensive system for clinical diagnostics, there has been the need to converge expertise in molecular biology and engineering.The goal remains to design miniaturised laboratory components that can be produced economicallywith superior performances with respect the macroscale systems.

Microelectromechanical systems (MEMS) are one potential solution because they leverage the economies of scale realised in the silicon processing industry, while permitting the exploration of new principles of sensing not otherwise possible with conventional methods.

One potential advantage of MEMS devices is their cost of manufacture: batch fabrication enables the production ofthousand of devices at a cost that is only incrementally higher than the cost of manufacturing single one. In some case MEMS based biomedical microsystems (MST-BioMEMS) are considered disposable, much like the plastics and biomedical reagents used in most laboratories. 

From draft design to an optimised microsystem module extensive work is required. Separate development and optimisation of the micromachined elements is an important advantage of the hybrid integration technique. Nowadays various types ofconstructive elements like grooves and channels as well as sensors have been realised . In addition, the use of silicon offers the unique possibility to realise specific transducers with standard microelectronic technologies.

 

 

Objectives

In this contest, the project main goal consists of providing new powerful micro-analytical microsystems, intended for the clinicaldiagnostic and able to perform molecular micro-analyses, based on functionalised microsensors integrated withmicrofabricated dispensers for on line micro-litres sample handling.

The proposal main objective has been focused on a special class of biomedical microsystems, based on multiple transducers elements, typically known as bio-chips. In our contest, a biochip is intended as a integrated microsystem used for biomedical assays that has an array of functionalised microtransducers and microprobes combined with well refined microfluidic modules. The integrated mycrosystems proposed herein will be designed to be used in situations where it is desirable to quickly screen large amounts of bio-molecule samples. The aim is to avoid time consuming manual steps and to make analysis as cost-effective as possible. 

Usually, the analytes of interest, such as small biomolecules, are usually present as a minor component in a complex mixture. This means that discrimination of the analyte from potential interference is a critical step in the analysis procedure. In order to overcome this drawback the adopted approach in our research is to incorporate both functionalised microtranducers and separation steps. This strategy will be developed into the concept of a “total bio-chemical analysis microsystem” where the main idea is to realise microsystems for performing all the component stages of a complete analysis in an integrated and automated fashion. These stages will include sample pre-treatment, chemical reactions, analytical separation, analyte detection and data analysis. 

In order to use the BioMEMS microfabrication technology, at least two major issues will be addressed. 

The first issue which has to be developed regards the reliable detection, separation and functionalisation techniques to study the adsorption of certain proteins on their surface and the detection of food borne pathogens by immobilizing monoclonal antibodies specific to the pathogenic strain onto detectors.Secondly, although sample manipulation mechanisms are widely understood, analysis of real samples require more sophisticated modelling, design and fabrication methodologies of microstructures fully devoted to the microfluidics and sample dispensing. 

The rationale of the project can be detailed in two main tasks:

i.)Recent progresses in micro-fabrication technologies have made feasible the development of detection techniques, based on electrochemical micro-transducers, capable of stable and high-throughput transduction of biological signals. 

In these applications both the change in impedance of microfabricated metallic microelectrode and in the charge of CMOS field effect microtransducers upon binding of the target of biological species to specific ligands immobilised on the sensor surfaces, will be proposed as detection mechanisms. 

Fabrication of these devices at the microelectronic scale leads to the “on-chip” creation of multi-analyte sensors or microelectrode array. The devices can be adapted as multi-analyte biosensors by immobilising a range of biological molecules on the microtransducer array. A basic step for the approach to the surface functionalisation concerns the development of a patterning methodology directly applied to the sensor surfaces in order to promote the molecular adhesion on specific sites. The proposed research activity in this field, after a preliminary theoretical modeling of the fundamental interaction process of model molecular systems with surfaces, will be devoted to the study of the controlled termination of surfaces by means of organic molecules and supramolecular aggregates, “ad-hoc” synthesized and to the understanding of the parameters and mechanisms critical to the adhesion process ofbiomolecules on ordered and disordered surface. 

ii.)The growth of microfluidic applications for medical diagnostics and bio-chemical assays has created the need for fundamental building blocks from which systems can be constructed. Microfluidics networks are used to pattern biomolecules with high resolution on a variety of substrate. However, in many micro chemical analysis or synthesis systems, flow control is a crucial issue. In our aims the whole system will be designed for total biochemical analysis, where injection of organic fluids in capillaries by means of pumps allows the fluids titration. Tipically these analyses require a non-pulsed fluid pumping. Because of the sometimes extremely narrow channels that are included in such systems, conventional flow control by hydraulic pumping cannot always be applied because excessively high pressures are required to achieve a significant flow rate. The application area of the project system influences some of features of its microfluidic components. Thus, non-mechanical mechanism (valveless) for pumping fluids, based on the use an electric field to induce a travelling wave along the microchannels,will be investigated in this project. 

For this reason one of the possible solution is the use of electroosmotic pumps where the reduced dead volumes increases the analytical performances. The working principle of an electroosmotic microfluidic circuit depends, among others, on the following parameters: liquid speed, flow rate, hydrostatic pressure, electroosmotic pressure. The control of these parameters is highly correlated to the geometry of the capillaries, e.g. length and diameters. Controlled and optimised design and fabrication of the microfluidic parts of the system will be possible with the support of a correct mathematical modelling of the structures. Numerical and analytical simulation of single parameters and of the overall microfluidic circuit will be performed at different stages of the project. Initially to help in the design of the first run, then in order to optimise further runs.

 

 

Conclusions

Summarising, our research proposal is focussed on the implementation of microfabrication technologies for new total bio-chemical analysis microsystems based on microtransducer array and micromachined modules: to reach these goals we envisage a very successful strategy to tackle both the techniques for sensor functionalisation and the fabrication strategies for valveless sample handling microfluidic networks .

For the applications in biomedicine and biotechnology of the proposed microsystems, we emphasize the need of a multidisciplinary approach. The necessary competencies include BioMEMS Microsystems design and fabrication, microfluidics, sensor surface functionalisation, neural networks, interfaces electronics and system automation.

The present multidisciplinar project is highly innovative, highly challenging and along the lines identified by the European Commission in the framework of the Future Emerging Technologies Programme.