This article presents an original work contributing to the rational design of imprinted catalyst by molecular self-assembly toward predetermined high specificity. Assembling with p-nitrophenyl phosphate as the transition state analogue (TSA) of p-nitrophenyl acetate esterolysis and 1-vinylimidazole as the functional monomer, the imprinted catalyst was prepared. An increase in the amount of assembled monomer results in a higher activity of hydrolysis, which, however, does not lead to an improvement of specificity. The best specificity is shown at the optimal self-assembly (corresponding to a stoichiometric interaction of monomer-TSA). Higher or lower an amount of assembled monomer would lead to a dramatic decrease in this specificity. Related information indicates that these may be a result of increasing specific interaction between the TSA and binding sites, which make the catalyst capable of selectively recognizing the transition state and promoting the conversion from reactant to the transition state.