The influence of thermo-mechanical treatment on microstructure and mechanical property of Ti-25Nb-2Mo-4Sn alloy was systematically investigated by X-ray diffraction, transmission electron microscopy and tensile test. Results show that the high-temperature β phase cannot be fully stabilized and a large amount of α" martensites forms after a solution treatment at 800 °C for 1 h followed by water quenching, which is caused by the low content of β-stabilizers (Nb and Mo) in Ti-25Nb-2Mo-4Sn alloy. In this case, the alloy exhibits low yield stress. It is also found that even under the same aging treatment of 400 °C for 2 h, the aging products for the solution treated specimen and the cold-rolled specimen are β + ω and β+α, respectively. Cold rolling deformation induces a mass of dislocations and grain boundaries could suppress the formation of ω and alternatively promote α phase precipitation. Upon a cold rolling and aging at 475 °C for 15 min, superior mechanical properties are achieved in Ti-25Nb-2Mo-4Sn alloy, with yield strength of 1033 MPa and tensile strength of 1113 MPa, combining with a low elastic modulus of 65 GPa.