Fiber-Optic Evanescent Wave Biosensor for the Detection of Oligonucleotides
- 1 September 1996
- journal article
- research article
- Published by American Chemical Society (ACS) in Analytical Chemistry
- Vol. 68 (17), 2905-2912
- https://doi.org/10.1021/ac960071+
Abstract
An automated optical biosensor system based on fluorescence excitation and detection in the evanescent field of a quartz fiber was used to detect 16-mer oligonucleotides in DNA hybridization assays. A biotinylated capture probe was immobilized on the fiber surface via avidin or streptavidin. The hybridization with fluorescein-labeled complementary strands was monitored in real time by fluorescence detection. The double strands formed by hybridization could be dissociated by chemical or thermal regeneration, allowing one to perform hundreds of assay cycles with the same fiber. The signal loss during long-time measurements, i.e., consecutive hybridization assays, can be described by a single-exponential function. Over more than 200 cycles, the net signal decreased by 50% with a signal variation of 2.4% after correction for this signal loss. By binding the capture probe with the 5‘-end to the optical fiber surface, and by using a 50% (w/w) aqueous urea solution for chemical regeneration, the duration of an assay cycle could be reduced to 3 min. By applying longer assay cycles, the detection limit for the hybridization with a complementary fluorescein-labeled oligonucleotide was 2.0 × 10-13 M (24 fmol). To detect an unlabeled complementary 16-mer oligonucleotide, competitive hybridization assays were performed, resulting in a detection limit of 1.1 × 10-9 M (132 pmol). Poly(acrylic acid) 5100 sodium salt and Tween 20 were used in the hybridization buffer to prevent nonspecific binding caused by ionic or hydrophobic interaction. The amount of nonspecific binding of noncomplementary oligonucleotides was in the range of 1−2%, compared with the specific binding in the different hybridization assays.Keywords
This publication has 16 references indexed in Scilit:
- Fiber-Optic DNA Sensor for Fluorometric Nucleic Acid DeterminationAnalytical Chemistry, 1995
- Detection of Aromatic Compounds Based on DNA Intercalation Using an Evanescent Wave BiosensorAnalytical Chemistry, 1995
- Sequence-Specific Gene Detection with a Gold Electrode Modified with DNA Probes and an Electrochemically Active DyeAnalytical Chemistry, 1994
- Voltammetric DNA Biosensor for Cystic Fibrosis Based on a Modified Carbon Paste ElectrodeAnalytical Chemistry, 1994
- Detection of Chlamydia trachomatis in clinical specimens Comparison of culture, direct antigen detection, DNA probe hybridization and PCRJournal of Microbiological Methods, 1994
- Grating couplers as label-free biochemical waveguide sensorsBiosensors and Bioelectronics, 1993
- Fiber-optic Atrazine immunosensorSensors and Actuators B: Chemical, 1993
- Gene probe assays on a fibre-optic evanescent wave biosensorBiosensors and Bioelectronics, 1992
- Hybridization of nucleic acids immobilized on a quartz crystal microbalanceJournal of the American Chemical Society, 1992
- Analytical methodology for immunoassays and DNA hybridization assays — current status and selected systems — critical reviewClinica Chimica Acta; International Journal of Clinical Chemistry, 1990