Development of surface plasmon resonance immunosensor for the novel protein immunostimulating factor

Abstract

The immunostimulating factor (ISTF) plays an important role in immunopathologic changes associated with Actinobacillus actinomycetemcomitans where it is located in the outer cell membrane. We describe a novel method by which ISTF can be monitored using a protein chip system based on surface plasmon resonance (SPR). The affinity of ISTF to its monoclonal antibodies and other proteins was compared. In fabrication of an ISTF immunosensor, a calix[4]crown ether monolayer was anchored onto a gold surface for use as an artificial protein linker system, and then characterized by Fourier Transform infrared reflection absorption spectroscopy, atomic force microscopy, and cyclic voltammetry. Anti-ISTF was immobilized onto the monolayer, and the interaction between ISTF and its antibodies was investigated by SPR. The calix[4]crown ether was found to assemble as a monolayer on the gold surface. Orientation and accessibility of anti-ISTF were assessed by the selective binding of ISTF. Modification of the SPR chip with the calix[4]crown monolayer provides a reliable and simple experimental platform for investigation of isolated proteins under experimental conditions resembling those of their native environment.