Evaluation of surface-enhanced resonance Raman scattering for quantitative DNA analysis.
The labeling of biological species using dyes has become common practice to aid in their detection, and immediate positive identification of specific dyes in high dilution is a key requirement. Here the detection by surface-enhanced resonance Raman scattering (SERRS) of eight commercially available dye labels (ROX, rhodamine 6G, HEX, FAM, TET, Cy3, Cy5, TAMRA) attached to oligonucleotide strands is reported. Each of the eight labels was easily detected by using the SERRS from silver nanoparticles to produce a unique, molecularly specific spectrum. The conditions were optimized to obtain the best signal enhancement, and linear concentration graphs at low oligonucleotide concentrations were obtained. At higher concentrations (above approximately 10(-)(8) mol dm(-)(3)), curvature was introduced into the concentration graphs with the exception of rhodamine 6G, TET, and FAM, which gave linearity over the entire concentration range studied. Detection limits as low as 0.5 fmol were obtained, with lower possible if a smaller sample was analyzed. Investigation was also carried out into the effect of a Tris-HCl buffer containing the surfactant Tween 20 to aid in the prevention of surface adhesion of the oligonucleotides to the sample vessels at ultralow concentrations. The Tween 20 allowed lower detection limits to be obtained for each of the labels studied. This study shows that the different dyes commonly used with oligonucleotides can give quantitative SERRS at concentration levels not possible when the same dyes are used with fluorescence detection.
Faulds
Karen
K
Department of Pure and Applied Chemistry, University of Strathclyde, 295 Cathedral Street, Glasgow, G1 1XL, UK.
Smith
W Ewen
WE
Graham
Duncan
D
D001665:Binding Sites; D002021:Buffers; D019343:Citric Acid / Q000737:chemistry; D003102:Colloids / Q000737:chemistry; D004247:DNA / Q000032:analysis* / Q000737:chemistry; D015342:DNA Probes / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D015394:Molecular Structure; D011136:Polysorbates / Q000737:chemistry; D012834:Silver / Q000737:chemistry; D013059:Spectrum Analysis, Raman / Q000379:methods*; D013096:Spermine / Q000737:chemistry; D013499:Surface Properties
D002021:Buffers; D003102:Colloids; D015342:DNA Probes; D005456:Fluorescent Dyes; D011136:Polysorbates; D019343:Citric Acid; D013096:Spermine; D012834:Silver; D004247:DNA