| Summary: | A DNA micro-optode for dengue virus detection was developed based on the sandwich hybridization strategy of DNAs on succinimide-functionalized poly(<i>n</i>-butyl acrylate) (poly(<i>n</i>BA-NAS)) microspheres. Gold nanoparticles (AuNPs) with an average diameter of ~20 nm were synthesized using a centrifugation-based method and adsorbed on the submicrometer-sized polyelectrolyte-coated poly(styrene-<i>co</i>-acrylic acid) (PSA) latex particles via an electrostatic method. The AuNP−latex spheres were attached to the thiolated reporter probe (rDNA) by Au−thiol binding to functionalize as an optical gold−latex−rDNA label. The one-step sandwich hybridization recognition involved a pair of a DNA probe, i.e., capture probe (pDNA), and AuNP−PSA reporter label that flanked the target DNA (complementary DNA (cDNA)). The concentration of dengue virus cDNA was optically transduced by immobilized AuNP−PSA−rDNA conjugates as the DNA micro-optode exhibited a violet hue upon the DNA sandwich hybridization reaction, which could be monitored by a fiber-optic reflectance spectrophotometer at 637 nm. The optical genosensor showed a linear reflectance response over a wide cDNA concentration range from 1.0 × 10<sup>−21</sup> M to 1.0 × 10<sup>−12</sup> M cDNA (<i>R<sup>2 </sup></i>= 0.9807) with a limit of detection (LOD) of 1 × 10<sup>−29</sup> M. The DNA biosensor was reusable for three consecutive applications after regeneration with mild sodium hydroxide. The sandwich-type optical biosensor was well validated with a molecular reverse transcription polymerase chain reaction (RT-PCR) technique for screening of dengue virus in clinical samples, e.g., serum, urine, and saliva from dengue virus-infected patients under informed consent.
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