Authors: G. Jia, K.S. Ma, H. Kido, J.V. Zoval and M.J. Madou
Affilation: University of California at Irvine, United States
Pages: 624 - 627
Keywords: flow-through, DNA immobilization, DNA hybridization, microfluidics, CD platform
A CD-based flow-through approach for DNA immobilization on gold surfaces was proposed and validated using 25-mer thiol-derivatized nucleotides (T-SH) labeled with fluorescent (Cy3) dye. Amino-derivatized ssDNA (T-amino) of the same sequence was used as a control. Passive immobilization was also conducted for comparison purposes. Three concentrations (1, 2.5, and 5 µM) of probe solutions were used in both flow-through and passive immobilization experiments. With the same immobilization time (10 minutes), the flow-through method resulted in significantly higher signal intensities at lower concentrations compared to the passive assay, as shown in Figure 3(a) & 3(b). This indicates that immobilization of thiol-derivatized DNA probes on gold can be accelerated using the proposed approach. Flow-through hybridization was performed using Cy3-labeled ssDNA complimentary to the sequence of the probes (without fluorescent labels). Our preliminary results in Figure 3(c) with a sample concentration of 100 nM have demonstrated not only the feasibility of the fast immobilization of DNA probes, but the potential of significant improvement in the limit of detection. Currently we are working on integrating both the DNA immobilization and the hybridization assay in a single microfluidic platform (flow cell) so that the total assay time may be reduced to less than 30 minutes.