Technical Abstract: A magnetic capture-based, surface-enhanced Raman scattering (SERS) assay for DNA detection has been developed which utilizes Au-coated paramagnetic nanoparticles (Au@PMPs) as both a SERS substrate and effective bio-separation reagent for the selective removal of target DNAs from solution. Hybridization reactions contained two target DNAs, sequence complementary reporter probes conjugated with spectrally distinct Raman dyes distinct for each target, and Au@PMPs conjugated with sequence complementary capture probes. In this case, target DNAs were derived from the RNA genomes of the Rift Valley Fever virus (RVFV) or West Nile virus (WNV). The hybridization reactions were incubated for a short period and then concentrated within the focus beam of an interrogating laser by magnetic pull-down. The attendant SERS response of each individually captured DNA provided a limit of detection sensitivity in the range of 20-100 nM. X-ray diffraction and UV-vis analysis validated both the desired surface plasmon resonance properties and bi-metallic composition of synthesized Au@PMPs, and UV-vis spectroscopy confirmed conjugation of the Raman dye compounds malachite green (MG)and erythrosin B (EB) with the RVFV and WNV reporter probes, respectively. Finally, hybridization reactions assembled for multiplexed detection of both targets yielded mixed MG/EB spectra and clearly differentiated peaks which facilitate the quantitative detection of each DNA target. Based on the simple design of a single-particle DNA detection assay, the opportunity is provided to develop magnetic capture-based SERS assays that are easily assembled and adapted for high-level multiplex detection using low-cost Raman instrumentation.