Research article

Oligodeoxyribonucleotide probe accessibility on a three-dimensional DNA microarray surface and the effect of hybridization time on the accuracy of expression ratios

David R Dorris^1,2 , Allen Nguyen^1,3 , Linn Gieser¹ , Randall Lockner^1,4 , Anna Lublinsky¹ , Marcus Patterson¹ , Edward Touma^1,4 , Timothy J Sendera^1,4 , Robert Elghanian^1,5 and Abhijit Mazumder^1,6

¹  Motorola Life Sciences, Northbrook, IL 60062, USA

²  Present address: Ambion, Austin, TX

³  Present address: Affymetrix, Palo Alto, CA

⁴  Present address: Amersham Biosciences, Tempe, AZ

⁵  Present address: Nanoink, Chicago, IL

⁶  Present address: Advanced Diagnostic Systems, Johnson and Johnson, Raritan, NJ

author email corresponding author email

BMC Biotechnology 2003, 3:6doi:10.1186/1472-6750-3-6

Published:	11 June 2003

Abstract

Background

DNA microarrays are now routinely used to monitor the transcript levels of thousands of genes simultaneously. However, the array fabrication method, hybridization conditions, and oligodeoxyribonucleotide probe length can impact the performance of a DNA microarray platform.

Results

We demonstrate solution-phase hybridization behavior of probe:target interactions by showing a strong correlation between the effect of mismatches in probes attached to a three dimensional matrix of a microarray and solution-based, thermodynamic duplex melting studies. The effects of mismatches in the probes attached to the microarray also demonstrate that most, if not all, of the oligodeoxyribonucleotide is available for hybridization. Kinetic parameters were also investigated. As anticipated, hybridization signals increased in a transcript concentration-dependent manner, and mismatch specificity increased with hybridization time. Unexpectedly, hybridization time increased the accuracy of fold changes by relieving the compression observed in expression ratios, and this effect may be more dramatic for larger fold changes.

Conclusions

Taken together, these studies demonstrate that a three-dimensional surface may enable use of shorter oligodeoxyribonucleotide probes and that hybridization time may be critical in improving the accuracy of microarray data.