Entwicklung und Charakterisierung von siliziumbasierten mikrotechnischen Komponenten für miniaturisierte DNA-Analysen

Abstract

This thesis deals with the design, fabrication and characterization of microsystems for use as single components within a miniaturized multistage DNA analysis. Within the first part of this work the example of a new microreaction chamber is used to demonstrate the suitability of silicon-based microsystems for DNA amplification by the temperature- and enzymatically controlled polymerase chain reaction (PCR). Covering design and fabrication aspects, two alternative system concepts are presented and compared. By the selected microchamber with double-sided heaters and sensor elements for temperature-controlling a high temperature homogeneity as well as maximum heating and cooling rates are achieved. A main focus of the characterization is the analysis of binding reactions of single-stranded oligonucleotide primers, double-stranded template molecules and the polymerase on microchamber surfaces. The successful DNA amplification in microchambers with volumes of 6 µl are demonstrated, whereas for a complete PCR with 30 cycles less than 18 min are needed.The second part is focused on a novel measurement principle for real-time detection of hybridization reactions on planar sensor surfaces by an impedance analysis of metallicinterdigitated electrodes. By using functionalized polymer microparticles, so-called microbeads, for labeling of DNA to be analyzed an increasing of the signal intensity is achieved.For modeling the sensor behaviour a two-dimensional electrical equivalent circuit is developed and applied for deriving of design and measurement criterions. The fabricated single sensor element consists of a thin film interdigitated electrode based on a tungsten titanium alloy with a dielectric passivation layer. Monocrystalline silicon and amorphous quartz glass are used as substrate materials. By experiments in a stopped-flow hybridization buffer the proof of functionality of the sensor element is provided.