Based on monolayer dispersion theory, Co₃O₄/ZSM-5 catalysts with different loadings have been prepared for selective catalytic reduction of nitrogen oxides by ammonia. Co₃O₄ can spontaneously disperse on HZSM-5 support with a monolayer dispersion threshold of 0.061 mmol 100 m^–2, equaling to a weight percentage around 4.5%. It has been revealed that the quantities of surface active oxygen (O₂^–) and acid sites are crucial for the reaction, which can adsorb and activate NO_x and NH₃ reactants effectively. Below the monolayer dispersion threshold, Co₃O₄ is finely dispersed as sub-monolayers or monolayers and in an amorphous state, which is favorable to generate the two kinds of active sites, hence promoting the performance of ammonia selective catalytic reduction of nitrogen oxide. However, the formation of crystalline Co₃O₄ above the capacity is harmful to the reaction performance. 4% Co₃O₄/ZSM-5, the catalyst close to the monolayer dispersion capacity, possesses the most abundant active O₂^– species and acidic sites, thereby demonstrating the best reaction performance in all the samples. It is proposed the optimal Co₃O₄/ZSM-5 catalyst can be prepared by loading the capacity amount of Co₃O₄ onto HZSM-5 support.