This paper presents the design, fabrication, and testing of a novel single stack-based piezoelectric energy harvester (PEH) for harvesting energy from three-degree-of-freedom (3-DOF) force excitation. One uniqueness lies in that the 3-DOF energy harvesting is implemented by using one piezoelectric stack. To scavenge energy from the 3-DOF input force, the proposed PEH is constructed with several force transmission mechanisms and slider mechanisms. The direction of the input force is first changed by the force transmission mechanisms, and the redundant force components are eliminated by the slider mechanisms. The transmitted force is then amplified by a two-stage force amplifier mechanism to improve the electric power output. The key parameters were found by establishing an analytical model of the proposed PEH. The best output performance of the PEH is achieved by selecting and optimally designing the key parameters. A prototype harvester was fabricated, and several experimental studies were conducted to verify the device performance. Results show the effectiveness of the developed 3-DOF PEH under the input force applied in x-axis, y-axis or z-axis. Furthermore, the issues that affect the practical application are discussed.