Adsorption-desorption of antigen to polystyrene plates used in ELISA

Since ELISA reliability depends to a great extent upon solid-phase reagent concentration and stability, we sought to analyse the influence of experimental conditions during ELISA performance on the adsorption/desorption of proteins to microplates. The effect upon desorption of several experimental parameters (antigen concentration, antibody concentration and affinity, washings, conjugate and inhibitor incubations) and quantitative treatment of protein-polystyrene adsorption were analysed. The adsorption to polystyrene microplates was studied with a hapten-conjugated protein (BSA-Ar36) in order to facilitate the analysis of the influence of antibody affinity on desorption during ELISA. Our results show that polystyrene plates adsorb BSA-Ar36 according to the Langmuir isotherm. The adsorption constant was 2.1 X 10(8) L/mol and maximal surface concentration of protein on solid phase was 1.8 X 10(-7) g/cm2. Although desorption was present, we observed that it did not affect the reliability of results of either direct or inhibition ELISA, because it was not dependent on the composition of the sample.