The efficient heat transfer with low pressure loss has long been a critical challenge in the microchannel heat sinks (MCHS) design. Further study is required for the quantitative analysis of the longitudinal vortex heat transfer enhancement mechanism. In this research, a microchannel heat transfer experiment and corresponding simulations are conducted. Based on the common Reynolds number range in engineering applications of MCHS, the underlying causes of heat transfer enhancement and pressure loss increase in MCHS with respect to Reynolds numbers is clarified. The results show that when the Reynolds number is 400, the cylindrical wake reduces the synergistic angle between the velocity and the temperature gradient at the streamwise gap, which greatly enhances the heat transfer performance of MCHS. In addition, when the Reynolds number is 900, the early separation of the cylindrical wake vortex leads to a significant reflux near the center of the spanwise gap, which significantly increases the pressure loss. A Different Reynolds number Efficiency Evaluation Criterion (DEEC) is proposed for evaluating the performance of MCHS. It is found that the flow state of cylindrical wake with Reynolds number of 400 has the best heat transfer efficiency. The concept of introducing longitudinal vortices into the pin–fin array can serve as a key guideline in the heat transfer enhancement optimization design of MCHS.