In situ grafting on surface was developed to modify porous microspheres. Functional monomers were grafted onto the porous microspheres. The grafting yield was up to 54.6%. The self-assembly of functional monomer and L-tryptophan molecules was verified by UV spectrometry. The surface imprinted polymer microspheres (SMIPs) were synthesized based on the in situ grafting on surface. The competitive binding experiments were carried out to assess the recognition. The results showed good selectivity of the SMIPs towards target molecule. Langmuir and Freundlich adsorption models were applied to description of the experimental adsorption isotherm. The analysis suggested that the adsorbing behavior of SMIPs was in agreement with Langmuir isotherm equation.