Sensitivity enhancement in DNA hybridization assay using gold nanoparticle-labeled two reporting probes.
A simple and sensitive method for DNA detection using gold nanoparticle (AuNP) two-probe detection system (AuNP-TP) was developed. Preliminary experiment was carried out by optimizing slide types, blocking agents and hybridization times. Fluorescent-labeled probes were used along with AuNP-labeled probes to confirm specific binding event between target DNA and probes. The sensitivities between AuNP single-probe (AuNP-SP) and AuNP-TP systems using sandwich-typed assay were compared. The AuNP-TP on epoxide-coated (EP) slides increased sensitivity 1000-fold at the detection limit of 100fM when compared to the AuNP-SP. This result indicates that the assay sensitivity was simply enhanced by simultaneous adding two AuNP labeled probes which selectively recognize different regions of the target DNA. The concept of AuNP-TP could potentially be applied to a macroarray format to detect multiple DNA targets simultaneously; thereby making the assay becomes more affordable and more sensitive.
Oaew
Sukunya
S
Biochemical Engineering and Pilot Plant Research and Development Unit, National Center for Genetic Engineering and Biotechnology, National Sciences and Technology Development Agency at King Mongkut's University of Technology Thonburi (Bangkhuntien), Bangkhuntien, Bangkok, Thailand. sukunya.oae@biotec.or.th
Karoonuthaisiri
Nitsara
N
Surareungchai
Werasak
W
D015374:Biosensing Techniques / Q000295:instrumentation*; D015342:DNA Probes / Q000737:chemistry* / Q000235:genetics*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D017930:Genes, Reporter / Q000235:genetics; D006046:Gold / Q000737:chemistry*; D017403:In Situ Hybridization / Q000295:instrumentation*; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000295:instrumentation*; D013194:Staining and Labeling
D015342:DNA Probes; D006046:Gold