Reservoir heterogeneity is regarded as one of the main reasons leading to low oil recovery for both conventional and unconventional reservoirs. High-permeability layers or fractures could result in ineffective water or gas injection and generate nonuniform profile. Polymer microspheres have been widely applied for the conformance control to overcome the bypass of injected fluids and improve the sweep efficiency. For the purpose of examining the plugging performance of submicron-sized microspheres in high-permeability porous media, systematic investigations were implemented incorporating macroscale blocking rate tests using core samples and pore-scale water migration analysis via nuclear magnetic resonance (NMR). Experimental results indicate that microsphere particle size dominates the plugging performance among three studied factors and core permeability has the least influence on the plugging performance. Subsequently, microsphere flooding was conducted to investigate its oil recovery capability. Different oil recovery behaviors were observed for cores with different permeability. For cores with lower permeability, oil recovery increased stepwise with microsphere injection whereas for higher permeability cores oil recovery rapidly increased and reached a plateau. This experimental work provides a better understanding on the plugging behavior of microspheres and could be employed as a reference for screening and optimizing the microsphere flooding process for profile control in heterogeneous reservoirs.