We report the first entanglement-based quantum key distribution (QKD) experiment over a 100-km optical fiber. We used superconducting single photon detectors based on NbN nanowires that provide high-speed single photon detection for the 1.5-mum telecom band, an efficient entangled photon pair source that consists of a fiber coupled periodically poled lithium niobate waveguide and ultra low loss filters, and planar lightwave circuit Mach-Zehnder interferometers (MZIs) with ultra stable operation. These characteristics enabled us to perform an entanglement-based QKD experiment over a 100-km optical fiber. In the experiment, which lasted approximately 8 hours, we successfully generated a 16 kbit sifted key with a quantum bit error rate of 6.9 % at a rate of 0.59 bits per second, from which we were able to distill a 3.9 kbit secure key.