Novel assay format for proteins based on magnetic molecularly imprinted polymer nanoparticles—detection of pepsin

AbstractA novel, abiotic assay format for the quantification of pepsin has been developed featuring replacement of conventional antibody binders and enzyme reporters with entirely synthetic alternatives. The assay was based on the application of magnetic molecularly imprinted polymer nanoparticles (MIP NPs) in an enzyme-linked immunosorbent assay format, where colored or fluorescent polystyrene beads serve as a label instead of enzyme. Modification of the standard microtiter plate with magnetic inserts permits the instantaneous immobilization of magnetic MIPs, with a hole in the middle of each insert allowing unobstructed measurement of the sample. The magnetic MIP NPs were synthetized using a solid-phase synthesis method in which pepsin, which was chosen as a model protein target present in “gastric juice”, was used as a template. The results suggested that blue polystyrene particles with 10 μm diameter combined with magnetic MIP NPs and magnetic inserts can be used for reliable detection of pepsin in th...