Applications in a variety of fields rely on the high-throughput ultrasensitive and multiplexed detection of oligonucleotides. However, the conventional microarray-based techniques that employ fluorescent dyes are hampered by several limitations; they require target amplification, fluorophore labeling, and complicated instrumentation, while the fluorophore-labeled species themselves exhibit slow binding kinetics, photo-bleaching effects, and overlapping spectral profiles. Among the emerging nanomaterials that are being used to solve these problems, oligonucleotide-gold nanoparticle conjugates (Oligo-Au NPs) have recently been highlighted due to their unique chemical and physical properties. In this chapter, a detection scheme for oligonucleotides that utilize Oligo-Au NPs is evaluated with multiple oligonucleotide targets. This scheme takes advantage of the sharp melting transitions, intense optical properties, catalytic properties, enhanced binding properties, and the programmable assembly/disassembly of Oligo-Au NPs.