This thesis reports design, fabrication, and implementation of two different micro electrophoresis system architectures for DNA analyses. The first architecture is traditional single channel layout with several design alternatives for size-based separation of DNA fragments. The second one is novel double channel architecture specialized for rapid mutation detection using heteroduplex analysis (HDA) method with an application of a newly designed injection technique. Besides achieving high resolution separations within the length of 1 mm with single channel arrangement, HDA was successfully applied for 590 base pair (bp) long PCR sample with 3 bp mutations in a separation length of 50 &micro / m in less than 3 minutes with double channel structure. Microchannels were formed using parylene-C due to its conformal deposition, no surface treatment requirement, transparency, biocompatibility, low background fluorescence, etc. Using the advantage of parylene in fabrication, the microchannels were fabricated with an only three-mask process. New double channel architecture is obtained by dividing the 200 &micro / m-wide separation channel into two parts by a 20 &micro / m-thick wall between them. For sample injection, various techniques, such as traditional cross, double-T, and double-L were investigated and optimized for single channel architecture assisting with pullback injection method. For double channel architecture, a novel, u-turn injection technique was applied. Precise control of sample amount by adjusting the injection time was accomplished by this new technique. Using high resolution cross-linked polyacrylamide gel as sieving material, separations were achieved in a very short length and time. Electrophoresis was performed in both channels of the double channel microchips simultaneously under the same conditions. This gives the chance of having a control channel in microchip format, which is very critical for the accuracy and reliability of the results in genetic analyses.

Thesis (Ph. D.)--University of Washington, 2000. / Vita. Includes bibliographical references (leaves 94-118).

Pancreatic cancer (PC) is extremely severe malignant disease with a five-year survival of less than 5%. Currently there is no reliable tool for the diagnosis of PC in its early stages. At the time of clinical symptoms most patients are in an advanced stage of the disease and the treatment does not usually have a significant effect. For these reasons emphasis is gradually shifting to the search for the suitable molecular markers for improvement of the diagnosis and assessment of the survival prognosis with respect to a possibility of surgical treatment. MiRNA represent one of the most promising markers, although, their examination in pancreatic tissue is a complicated process. One of the reasons is the very small amount of the source material coming from a fine needle biopsy. A second cause of problems is the subtle character of the pancreatic tissue resulting in significantly lower yields of molecular genetic analysis when compared to other epithelial tissues. An additional negative factor is heterogeneity of the tissue resulting in disproportionate representation of tumor cells within the sample. A suitable choice of procedures for isolation of nucleic acids (NA) and subsequent analysis including quantification of tumor cells is critical for accurate evaluation of the miRNA levels. This work is...