Detection of DNA Hybridization via Fluorescence Intensity Variations of ZnSe-DNA Quantum Dot Biosensors

Abstract

ZnSe quantum dots (QDs) that were capped with 11-mercaptoundecanoic acid (MUA) and conjugated to amino-modified ssDNA molecules exhibited variations in fluorescence emission intensity upon hybridization with complementary ssDNA in solution, a phenomenon that can be exploited for rapid detection of free ssDNA sequences. Conjugation of MUA-capped ZnSe QDs to amino-modified ssDNA molecules resulted in increased fluorescence emission intensity and stability at room temperature. Increasing the length of the ssDNA, that was conjugated to the QDs, resulted in increased fluorescence emission intensity up to a length of about 50 nucleotide bases, beyond which the peak emission intensity reached a plateau. Hybridization of QD-ssDNA conjugates with complementary ssDNA, either in free form or bound to QDs from the same population, resulted in additional fluorescence emission intensity amplification. A small red shift was observed when three-dimensional QD-dsDNA-QD structures were formed. The QD-ssDNA sensors with single ssDNA molecule per QD were developed and used for rapid quantitative detection of fully or partially complementary free ssDNA sequences in aqueous solution. Partial hybridization of the QD-ssDNA sensors with short ssDNA targets resulted in smaller QD emission intensity amplification, when compared to full hybridization. A QD-ssDNA sensor containing a sequence corresponding to the hemoglobin beta gene was used to detect and discriminate between free ssDNA targets consisting of a complementary ssDNA sequence and targets containing a single-base mutation that can cause sickle-cell anemia. Such QD-based biosensors can form the basis for rapid separation-free assays that can be used to detect target biomolecules in solution.

Keywords:

• Biosensors
• DNA mutation detection
• DNA sequence detection
• Flourescence intensity variation
• Homogeneous separation-free assays
• Nanocrystal-DNA bioconjunction
• Quantum dots
• Zinc selenide nanocrystals

Acknowledgments

This paper was submitted as part of a Special Issue on Biosensors organized by Dr. Yu Lei of the University of Connecticut.

The authors acknowledge financial support by the University of Massachusetts President's Science and Technology Fund (UMass NanoMedicine Institute), the NSF Center for Hierarchical Manufacturing (NSF-NSEC CMMI-0531171), the University at Buffalo IRCAF Multidisciplinary Research Fund, and the State of New York GSEU Professional Development Fund.