In this paper, a localized surface-plasmon resonance (LSPR) biosensor, which uses a U-shaped multi-mode fiber (U-MMF), is introduced and investigated. It is modified with a complex of three-dimensional (3D) gold nanoparticles and multilayer graphene as spacer: n*(Au/G)@U-MMF, where n denotes the layer number of gold nanoparticles. The gold nanoparticles were synthesized by reducing chloroauric acid. Graphene films were formed using a liquid/chemical method. The number of gold-nanoparticle layers was found to be critical for the performance of the sensor. Moreover, using the finite-difference time domain, 3D nanostructures, with a wide range of gold-nanoparticle layers, were explored. The sensor showed the sensitivity of 1251.44 nm/RIU, as well as high stability and repeatability; for the measurement-process of time- and concentration-dependent DNA hybridization kinetics with detection concentrations, ranging from 0.1nM to 100 nM, the sensor displayed excellent performance, which points towards a vast potential in the field of medical diagnostics.