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ARS Home » Plains Area » Manhattan, Kansas » Center for Grain and Animal Health Research » ABADRU » Research » Publications at this Location » Publication #236024
Title: SERS detection of indirect viral DNA capture using colloidal gold and methylene blue as a Raman label

Author
item Harpster, Mark
item ZHANG, HAO - UNIVERSITY OF WYOMING
item KUMAR, AJAYA - UNIVERSITY OF WYOMING
item RAY, BRYAN - DELTA NU
item WARD, TIMOTHY - UNIVERSITY OF WYOMING
item CARRON, KEITH - DELTA NU
item CORCORAN, ROBERT - UNIVERSITY OF WYOMING
item Mecham, James
item Wilson, William
item JOHNSON, PATRICK - UNIVERSITY OF WYOMING

Submitted to: Biosensors and Bioelectronics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/19/2009
Publication Date: 12/15/2009
Citation: Harpster, M.H., Zhang, H., Kumar, A., Ray, B.H., Ward, T., Carron, K.T., Corcoran, R.C., Mecham, J.O., Wilson, W.C., Johnson, P.A. 2009. SERS detection of indirect viral DNA capture using colloidal gold and methylene blue as a Raman label. Biosensors and Bioelectronics. 25(4):674. DOI: 10.1016/j.bios.2009.05.020.

Interpretive Summary: An indirect DNA capture model assay nanoparticles is demonstrated for using a Raman spectrophotometer detection system. The sequence targeted for capture is derived from the West Nile Virus (WNV) RNA genome and was selected based on computer analysis. The WNV target is captured associated to the nanopartical by a strong tethering group on the capture molecule and detected by a complementary reporter molecule is Raman active. The system was optimized and verified using Quartz Crystal Microbalance-Dissipation as a tool. The system is adapted to a compact Raman spectrometer, which is designed for analyte detection in capillary tubes, this assay provides a rapid, mobile and cost effective alternative to expensive spectroscopic instrumentation, which is often restricted to analytical laboratories.

Technical Abstract: An indirect capture model assay using colloidal Au nanoparticles is demonstrated for surface enhanced Raman scattering (SERS) spectroscopy detection of DNA. The sequence targeted for capture is derived from the West Nile Virus (WNV) RNA genome and was selected on the basis of exhibiting minimal secondary structure formation. Upon incubation with colloidal Au, hybridization of the WNV target sequence with a complementary capture oligonucleotide conjugated to a strong tethering group and a complementary reporter oligonucleotide conjugated to methylene blue (MB), a Raman label, anchors the resultant ternary complex to Au nanoparticles and positions MB within the required sensing distance for SERS enhancement. The subsequent elicitation of surface enhanced plasmon resonance by laser excitation provides a spectral peak signature profile that is capture-specific and characteristic of the Raman spectrum for MB. Detection sensitivity is in the sub-micromolar range and was shown to be highest for thiol, and less so for amino, modifications at the 5’ terminus of the capture oligonucleotide. Finally, using Quartz Crystal Microbalance-Dissipation as a tool for modeling ternary complex binding to Au surfaces, quantitative measurements of surface mass coverage on Au plated sensor crystals established a positive correlation between levels of ternary complex adsorption and their correspondent levels of SERS signal intensification. Adapted to a compact Raman spectrometer, which is designed for analyte detection in capillary tubes, this assay provides a rapid, mobile and cost effective alternative to expensive spectroscopic instrumentation, which is often restricted to analytical laboratories.