Published online 23 January 2004
Nucleic Acids Research, 2004, Vol. 32, No. 2 e18
© 2004 Oxford University Press
Rapid detection of single nucleotide polymorphisms associated with spinal muscular atrophy by use of a reusable fibre-optic biosensor
Chemical Sensors Group, University of Toronto at Mississauga, 3359 Mississauga Road North, Mississauga, Ontario, Canada L5L 1C6 and 1 Children’s Hospital of Eastern Ontario Research Institute, Molecular Genetics Laboratory, 401 Smyth Road, Ottawa, Ontario, Canada K1H 8L1
*To whom correspondence should be addressed. Tel: +1 905 828 5453; Fax: +1 905 828 5425; Email: ppiunno@utm.utoronto.ca
Rapid (<2 min) and quantitative genotyping for single nucleotide polymorphisms (SNPs) associated with spinal muscular atrophy was done using a reusable (approximately 80 cycles of application) fibre-optic biosensor over a clinically relevant range (0–4 gene copies). Sensors were functionalized with oligonucleotide probes immobilized at high density (
7 pmol/cm2) to impart enhanced selectivity for SNP discrimination and used in a total internal reflection fluorescence detection motif to detect 202 bp PCR amplicons from patient samples. Real-time detection may be done over a range of ionic strength conditions (0.1–1.0 M) without stringency rinsing to remove non-selectively bound materials and without loss of selectivity, permitting a means for facile sample preparation. By using the time-derivative of fluorescence intensity as the analytical parameter, linearity of response may be maintained while allowing for significant reductions in analysis time (10–100-fold), permitting for the completion of measurements in under 1 min.