FIBEROPTIC DNA SENSOR FOR FLUOROMETRIC NUCLEI ACID DETERMINATION

Single-stranded deoxyribonucleic acid (ssDNA) thymidylic acid icosanuc leotides (dT(20)) were synthesized on the surfaces of derivatized quar tz optical fibers to create an optical DNA biosensor. The synthesis ma de use of an automated solid-phase synthesizer and phosphoramidite syn thons, The covalently immobilized oligomers were found to hybridize wi th complementary ssDNA (cDNA) or ssRNA (cRNA) from solution, and the d evice was regenerable for multiple cycles of application. Hybridizatio n on optical fibers was detected by the use of the fluorescent DNA sta in ethidium bromide (EB). The procedure used hybridization assay techn iques and provided a detection limit of 86 ng . mL(-1) cDNA and a sens itivity of 200% fluorescence intensity increase per 100 ng . mL(-1) of cDNA, with one cycle of hybridization analysis requiring 45 min, The sensor has been observed to be regenerable (minimum of five cycles) an d to sustain full activity after prolonged storage times (1 year), har sh washing conditions (sonication), and sterilization (autoclaving). T he extent of hybridization between the immobilized and complementary n ucleic acid strands was determined by UV absorbance thermal denaturati on studies wherein ah 20 bases on each strand of the nucleic acid were found to be involved in duplex formation.