Planar waveguide biosensors for nucleic acid hybridization reactions

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Publication Type Journal Article
School or College College of Engineering
Department Bioengineering
Creator Christensen, Douglas A.
Other Author Herron, James N.; zumBrunnen, Stacy; Wang, Jian-xun; Gao, Xiao-lin; Wang, Hsu-kun; Terry, Alan H.
Title Planar waveguide biosensors for nucleic acid hybridization reactions
Date 2000
Description Oligonucleotide probes derived from (1) the T3 RNA polymerase promoter sequence (T3) and (2) prostate-specific antigen messenger RNA (PSA) were prepared and labeled with a red-emitting fluorescent dye (Cy5). The complimentary oligonucleotides (anti T3 and anti PSA) were prepared and labeled with biotin. Initially, a feasibility study was performed in which the hybridization rate of the T3/anti T3 oligonucleotide pair was examined. Specifically, biotinylated anti T3 was immobilized to a neutravidin-coated waveguide and solutions containing increasing concentrations of Cy5-labeled T3 were injected into the biosensor. Fluorescence emission was detected with an evanescent wave imaging fluorometer. The hybridization reaction proceeded rapidly with a significant amount of binding occurring during the first 5 minutes. A Michaelis-Menton kinetics model was used to analyze hybridization rate data and gave values of 78 nanomolar for the apparent affinity of the hybridization reaction and 1.4 picomolar for the analytical sensitivity of the hybridization assay. In subsequent studies the hybridization rate of the PSA/anti PSA oligonucleotide pair was examined. Biotinylated anti PSA was immobilized to the waveguide and solutions containing increasing concentrations of Cy5-labeled PSA were injected into the biosensor. The hybridization rate observed for formation of the PSA/anti PSA pair was comparable to the high rates observed for the T3/anti T3 pair. Lastly, the selectivity of the biosensor was examined using an oligonucleotide probe derived from human glandular kallikrein (hGK), which exhibits a high degree of homology to PSA. The two oligonucleotide probes (PSA and hGK) only differed in 7 out of 20 positions. Interestingly, the hybridization rate observed for Cy5-Iabeled hGK was very low-not statistically different from the non-specific binding rate of the hybridization assay.
Type Text
Publisher International Society for Optical Engineering (SPIE)
Volume 3913
First Page 177
Last Page 184
DOI 10.1117/12.382029
citatation_issn 0277786X
Subject Planar waveguides; Evanescence; Fluorescence; Glandular kallikrein; MDx; Molecular diagnostics; Nucleic acid; PSA; Oligonucleotide probes
Subject LCSH Biosensors; Wave guides; Nucleic acid hybridization; Molecular diagnosis; Prostate-specific antigen
Language eng
Conference Title In-Vitro Diagnostic Instrumentation; Wednesday 26 January 2000; San Jose, CA, USA
Bibliographic Citation Herron, J. N., zumBrunnen, S., Wang, J.-X., Gao, X.-L., Wang, H.-K., Terry, A. H., & Christensen, D. A. (2000). Planar waveguide biosensors for nucleic acid hybridization reactions. Proceedings of SPIE - The International Society for Optical Engineering, 3913, 177-84.
Rights Management (c)Society of Photo-Optical Instrumentation Engineers. One print or electronic copy may be made for personal use only. Systematic electronic or print reproduction and distribution, duplication of any material in this paper for a fee or for commercial purposes, or modification of the content of the paper are prohibited.
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Format Extent 656,803 bytes
Identifier ir-main,11665
ARK ark:/87278/s6w38dtc
Setname ir_uspace
Date Created 2012-06-13
Date Modified 2021-05-06
ID 706099
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