Visual detection of labeled oligonucleotides using visible-light-polymerization-based amplification.
DNA biochip technology holds potential for highly parallel, rapid, and sensitive genetic diagnostic screening of target pathogens and disease biomarkers. A primary limitation involves a simultaneous, sequence-specific identification of low copy number target polynucleotides using a clinically appropriate detection methodology that implements only inexpensive detection instrumentation. Here, a rapid (20 min), nonenzymatic method of signal amplification utilizing surface-initiated photopolymerization is presented in glass microarray format. Visible light photoinitiators covalently coupled to streptavidin were used to bind biotin-labeled capture sequences. Amplification was achieved through subsequent contact with a monomer solution and the appropriate light exposure to generate 20-240-nm-thick hydrogel layers exclusively from spots containing the biotin-labeled DNA. An amplification factor of 10(6) to 10(7) was observed as well as a detectable response generated from as low as approximately 10(4) labeled oligonucleotides using minimal instrumentation, such as an optical microscope or CCD camera.
Department of Chemical and Biological Engineering, ECCH 111 CB 424, University of Colorado, Boulder, Colorado 80309, USA.
Ryan R
RR
Hansen
Hadley D
HD
Sikes
Christopher N
CN
Bowman
D001483:Base Sequence; D017931:DNA Primers; D008027:Light; D009841:Oligonucleotides / Q000032:analysis*; D011108:Polymers / Q000737:chemistry*; D012680:Sensitivity and Specificity
D017931:DNA Primers; D009841:Oligonucleotides; D011108:Polymers