Objective To improve the measuring results of enzyme immunoassay (EIA) by appropriate modification of polystyrene microtitre plates (PS) with several different methods.Methods The interaction between enzyme and its substrate was investigated by using horseradish peroxidase (HRP) as immobilizing molecules.The coating antigen molecules,including recombinant human augmenter of liver regeneration (ALR),double strand DNA and cardiolipin,and the coating anti-ALR antibody molecules with different status (unoxidized and oxidized antibody) were immobilized on the modified PS plates respectively and investigated by EIA.A synchronous EIA was performed to compare the results by using untreated PS plates.Results Besides the feature of solid-supports,the immobilizing effect of biomolecules was strongly associated with their intrinsic characteristics and physical status,including the molecular classification and structures,molecular weight and electric charge on the molecular surface,etc.Among them,the surface modified by APTES and PLL improved significantly the solidifying effect of ALR,dsDNA,cardiolipin and oxidized antibody and enhanced the absorbance value and sensitivity of EIA.The EIA measurement using ultraviolet-irradiated plate for anti-ALR and anti-dsDNA antibody and aldehyde-surface modified by glutaraldehyde (GA) for anti-cardiolipin antibody presented higher specific signals and lower background noise.The results of atomic force microscopy (AFM) characterization demonstrated that adsorbed biomolecules based on hydrophobic interaction exhibited randomized and irregular distribution and clustered agglutination,whereas the dimensional tropism and performance of the molecules immobilized based on the manner of covalent bond and attractive linkage of electric charge evenly spread on solid supports and presented homogeneous distribution.The amount of molecules adsorbed on the surfaces and their function-preserving level were significantly increased.Conclusion The modified PS plates increase the capacity of immune molecules attachment.The satisfied performance and effectiveness of EIA on solid supports may be achieved by a given pattern in which covalent bond is combined with electric charge attraction.