A paramagnetic-reporter two-particle system for amplification-free detection of DNA in serum

Quantitative nucleic acid detection is used extensively in the management of many pathogenic infections, where viral or bacterial nucleic acid copy number relates directly to disease prognosis. Temperature-cycle or isothermal amplification formats offer excellent performance, but their requirement for purified nucleic acid and accurate temperature control increases costs and renders their migration to resource-limited environments problematic. In contrast, amplification-free nucleic acid assays could allow simplified system design, resulting in reduced costs. In this study, we report a amplification-free herpes simplex virus (HSV) assay where oligoethylene glycol methacrylate (OEGMA) grafted ssDNA capture-probes on paramagnetic nanoparticles are coupled with reporter-probe-modified fluorescent nanoparticles in a target-dependent manner. Following assay and reagent optimization, a sub-pM (25 amol) limit of detection could be achieved in buffer and also in neat, undiluted serum, representing a 160-fold improvement over that achieved using convention detection with a fluorescence plate reader. Equivalent performance in serum and buffer offers the opportunity for simplified diagnostic device design for resource-limited environments.