Abstract: 3D printing technology has huge advantages in rapid prototyping and fabricating parts with complex structures. Three microchannel heat sinks are fabricated by 3D printing process, which contain two open single-layer microchannels and one closed triple-layer microchannel. A pressure test is carried out for checking the mechanical property of the three-layered microchannel heat sink (TLMHS). The hydrodynamic and heat transfer characteristics of TLMHS are investigated by experiment and numerical simulation under the conditions of constant heat flux and Reynolds number ranging from 50 to 780. By comparing the experimental and numerical results, the influence of rough wall caused by 3D printing process on pressure drop and heat transfer is evaluated. The results show that the effect of rough wall on pressure drop is always negative, but that on heat transfer changes from negative to positive as Reynolds number increases. The heat transfer enhancement is quantitatively analyzed by defining the efficiency factors of energy and flow rate. It is found that the pump power is most efficient for heat transfer reinforcement when the Reynolds number is 244 and there is a pump power threshold that makes it unsustainable for heat transfer reinforcement.