To achieve true 3D nano-measurement with sub-nanometer resolution and very low touch force through a micro/nano coordinate measuring machine, a new 3D resonant trigger probe based on a quartz tuning fork is proposed. In this trigger probe, a quartz tuning fork with a microsphere tip vibrates at its resonant frequency, and is used as the sensing element. The resonance parameters of this quartz tuning fork (e.g., vibrating amplitude and resonant frequency) are extremely sensitive to external 3D microforces. The distinguished feature of this probe is its ability to interact with the sample surface in the actual three directions. The microsphere tip of the probe interacts with the sample surface in tapping mode in the Z direction, whereas it interacts in friction mode in the X and Y directions. The dynamic contact mechanism of the probe is based on interfacial force theory, and mechanical models of the interactions between the microsphere tip and sample surface in the X, Y, and Z directions are constructed and simulated. The experiment shows that the probe has sub-nanometer resolution in 3D directions and triggers repeatability of approximately 40 nm in each direction. Theoretical analysis and experimental results verify that this 3D resonant trigger probe can be used for true 3D profile measurement.