QUARTZ-CRYSTAL MICROBALANCE STUDY OF DNA IMMOBILIZATION AND HYBRIDIZATION FOR NUCLEIC-ACID SENSOR DEVELOPMENT

The immobilization of two 30-mer oligonucleotides, one biotinylated (b iotin-DNA) and the other having a mercaptohexyl group at the 5'-phosph ate end (BS1-SH), onto modified gold surfaces has been examined using a quartz crystal microbalance (QCM). Both single-layer and multilayer DNA films were prepared. The single-layer films to biotin-DNA were con structed by binding to a precursor layer of avidin, which had been att ached to the QCM either covalently using a water-soluble carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) or via electrostati c interaction with poly(allylamine hydrochloride) (PAH). Single-layer films of BS1-SH were also formed on PAH via the electrostatic attracti on between the amine groups on PAH and the negatively charged phosphat e backbone of DNA. Multilayer films of DNA were fabricated by the succ essive deposition of avidin and poly(styrenesulfonate) (PSS), up to a total of nine avidin/PSS layers, followed by DNA adsorption. DNA immob ilization and hybridization of the immobilized DNAs was monitored in s itu from QCM frequency changes. Hybridization was induced by exposure of the DNA-containing films to complementary DNA in solution. Equal fr equency changes were observed for the DNA immobilization and hybridiza tion steps for the single-layer films, indication a DNA probe-to-hybri dized DNA target ration of 1:1. The multilayer DNA films also exhibite d DNA hybridization, with a greater quantity of DNA hybridized compare d with the single-layer films. The multilayer films provide a novel me ans for the fabrication of DNA-based thin films with increased capacit y for nucleic acid detection.