Immunoassay utilizing biochemistry reaction product via surface-enhanced Raman scattering in near field

We propose here a kind of applications of surface-enhanced Raman scatterin (SERS) to immunology. It is a new enzyme immunoassay based on SERS. In the propose system, antibody immobilized on a solid substrate reacts with antigen, which binds with another antibody labeled with peroxidase. If this immunocomplex is subjected to reaction with o-phenylenediamine and hydrogenperoxide, azoaniline is generated. This azo compound is adsorbed on a silver colloid and only the azo compound gives a strong surface-enhanced resonance Raman (SERRS) spectrum. A linear relationship was observed between the peak intensity of the N=N stretching band and the concentration of antigen, revealing that one can determin the concentration of antigen by the SERRS measurement of the reaction product. The detectic limit of this SERS enzyme immunoassay method was found to be about 10^-15 mol/L.