Fischer-Tropsch is an important technology for the efficient use of energy such as coal, natural gas and biomass which can convert syngas into high value-added chemicals. Co-based catalysts are promising Fischer-Tropsch synthesis industrial catalysts due to their excellent carbon growth ability and low water vapor shift reaction. In order to improve the performance of Co-based catalysts, it is necessary to conduct a comprehensive and in-depth study and exploration of its Fischer-Tropsch synthesis mechanism. In this paper, the adsorption and activation of CO on the surface of cobalt-based catalysts were investigated by in- situ DRIFTS, in-situ Raman, in-situ XPS, XRD and chemical transient, adsorption and desorption methods. It was found that there are three adsorption modes of CO on the surface of the catalyst, such as line type, bridge type and hollow type, and bridge type adsorption is mainly found at room temperature. As the temperature increases, the bridge adsorption CO decreases rapidly, and the catalyst retains only linear adsorption above 200 °C. This shows that linear adsorption of CO has higher thermal stability. During the heating process in H2 atmosphere, there is a significant transition from the bridge-to-line morphology of the adsorbed CO. Only CO in the linear adsorption was observed on the surface of the catalyst at the reaction temperature, and the formation of C3H8 was apparent after the introduction of H2. Therefore, the linear adsorption was more likely to be an adsorption form having Fischer-Tropsch synthesis activity. This study can provide a reference for the exploration of cobalt-based Fischer-Tropsch synthesis mechanism.