Nanoparticle-Based Biobarcoded DNA Sensor for the Rapid Detection of pagA Gene of Bacillus Anthracis

Abstract

Bacillus anthracis is a bioterrorism agent classified by the Centers for Disease Control and Prevention (CDC). A highly amplified, nanoparticle-based, biobarcoded electrochemical biosensor for the rapid detection of pagA gene (accession number = M22589) in Bacillus anthracis is reported in this paper. The biosensor system is mainly composed of two nanoparticles: gold nanoparticles (AuNPs) and magnetic microparticles (MMPs). The AuNPs are coated with the 1st target-specific DNA probe (pDNA), which can recognize the target DNA (tDNA), and nanoparticle tracer (NT)-labeled barcode DNA (bDNA) as a signal indicator in a 1:100 probe-to-barcode ratio. The MMPs are coated with the 2nd target-specific pDNA. After mixing the nanoparticles with the tDNA, the sandwich structure (MMPs-2nd pDNA/tDNA/1st pDNA-AuNPs-bDNA-NTs) is formed. A magnetic field is applied to separate the sandwiches from the unreacted materials. Because the AuNPs have a large number of NTs per pDNA binding event, there is substantial amplification. Then, the NTs are dissolved in 1-M nitric acid and the metal ions (Pb <sup xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">2+</sup> and Cd <sup xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">2+</sup> ) are detected by square-wave anodic stripping voltammetry on a screen-printed carbon electrode sensor. Using this technique, the detection limit of this biobarcoded DNA sensor is as low as 0.2 ng/mL using cadmium sulfide NTs, or 0.02 ng/mL using lead sulfide NTs. The nanoparticle-based biobarcoded DNA sensor has potential applications in multiplexed detection of bioterrorism threat agents.