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Real-time detection of DNA hybridization and melting on oligonucleotide arrays by using optical wave guides

Stimpson, Don I. and Hoijer, Joanell V. and Hsieh, Wangting and Jou, Cynthia and Gordon, Julian and Theriault, Tomas and Gamble, Ronald and Baldeschwieler, John D. (1995) Real-time detection of DNA hybridization and melting on oligonucleotide arrays by using optical wave guides. Proceedings of the National Academy of Sciences of the United States of America, 92 (14). pp. 6379-6383. ISSN 0027-8424. PMCID PMC41521. doi:10.1073/pnas.92.14.6379.

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The challenge of the Human Genome Project is to increase the rate of DNA sequence acquisition by two orders of magnitude to complete sequencing of the human genome by the year 2000. The present work describes a rapid detection method using a two-dimensional optical wave guide that allows measurement of real-time binding or melting of a light-scattering label on a DNA array. A particulate label on the target DNA acts as a light-scattering source when illuminated by the evanescent wave of the wave guide and only the label bound to the surface generates a signal. Imaging/visual examination of the scattered light permits interrogation of the entire array simultaneously. Hybridization specificity is equivalent to that obtained with a conventional system using autoradiography. Wave guide melting curves are consistent with those obtained in the liquid phase and single-base discrimination is facile. Dilution experiments showed an apparent lower limit of detection at 0.4 nM oligonucleotide. This performance is comparable to the best currently known fluorescence-based systems. In addition, wave guide detection allows manipulation of hybridization stringency during detection and thereby reduces DNA chip complexity. It is anticipated that this methodology will provide a powerful tool for diagnostic applications that require rapid cost-effective detection of variations from known sequences.

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Additional Information:© 1995 by the National Academy of Sciences. Contributed by John D. Baldeschwieler, February 3, 1995. We are grateful to D. Zakula for the English translation of the abstract in ref. 1, M. McMahon for valuable discussions and pioneering work in the area of DNA hybridization diagnostics, T. Brainard for improvements to the manuscript, and W. Schultz for construction of the wave guide light source. The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Issue or Number:14
PubMed Central ID:PMC41521
Record Number:CaltechAUTHORS:STIpnas95
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Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:1412
Deposited By: Tony Diaz
Deposited On:18 Jan 2006
Last Modified:08 Nov 2021 19:10

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