Identificador: TDX:339
Autores: Pescador Álvarez, Paula
Resumen:
The layer-by-layer (LbL) coating of charged surfaces with oppositely charged materials is a powerful and versatile approach for the fabrication of functional molecular assemblies and interfaces. The key advantages of this technique over other methodologies such as self-assembled monolayers (SAM) or Langmuir-Blodgett (LB) are its unparalleled flexibility in combination with its simplicity and inexpensiveness. With simple instrumentation and easy preparation steps it is possible to assemble highly complex and stable architectures with nanoscale control over their composition and structure.The LbL approach is particularly suitable for the integration of biomolecules (proteins, lipids, DNA) into functional multilayers, since layer buildup is carried out under mild conditions. Many biologically relevant species can be incorporated into the films while maintaining or even improving their biological functions. A further advantage of this technique is that substrates of virtually any size and shape can be coated with functional films in a simple and controlled fashion. Of particular interest has been the extension of the LbL method to the modification of colloidal particles. Apart from their interest from a fundamental and applied point of view, colloids have emerged as powerful tools particularly suited to meet the challenge of creating tailored building blocks for the rapidly evolving fields of bio- and nanotechnology. The LbL functionalisation of colloidal particles provides a new route for the creation of composite architectures which allow the integration of nanoscale-defined materials into two- and three-dimensional structures. This ultimately opens the way not only to functional microsystems but also to the fabrication of macroscopic devices.One of the fields in which the LbL technique has represented a major advance is the development of electrochemical systems. Electrochemically active materials can be readily incorporated into multilayer films, together with other species which provide additional functionalities. Furthermore, critical parameters such as film thickness, mass transport and conductivity can be precisely tuned, allowing an increased control over the performance of the system. In particular, LbL assemblies have found many successful applications in the area of electrochemical biosensors. These devices provide an interface between biological functions and electronic signal-transduction processes, and offer great potential for the development of new miniaturised, low-cost, integrated biodetection platforms.In the present work, the LbL technique is employed to assemble multilayer films of enzymes and polyelectrolytes on the surface of silica microparticles. Two different enzymes, glucose oxidase (GOx) and horseradish peroxidase (HRP) are co-immobilised together with precursor and intermediate polyelectrolyte layers. In the resulting multilayer films a sequential reaction takes place, with the conversion of glucose to gluconic acid and hydrogen peroxide catalysed by GOx and the subsequent reduction of hydrogen peroxide to water catalysed by HRP. The sequential LbL approach allows to explore the influence of different polyelectrolyte combinations on the immobilisation and functionality of the enzymes. Flow cytometry, confocal and electron microscopy and spectrophotometric measurements provide information about the interaction between the different layer components, as well as the stability of the colloidal substrates and the behaviour of the multilayer films in the presence of the different enzyme substrates.An electrochemical functionality is further added to these films with the incorporation of an osmium-based redox polymer to the structure. In this way the specific chemical events taking place at the redox centers of the enzymes are transduced into an electrical signal. The nanostructured particles assume a multiple role in the final system. On one hand, they act as immobilisation substrates and high surface area carriers for the creation of enzymatic microreactors. Moreover, the LbL-coated colloids are embedded in a redox polymer film and immobilised on the surface of gold electrodes, acting as building blocks fo the fabrication of an electrochemical biosensor able to detect glucose and hydrogen peroxide.
Repositori URV