Immunoassays at a quartz-liquid interface: theory, instrumentation and preliminary application to the fluorescent immunoassay of human immunoglobulin G

The theoretical basis and instrumental requirements of an optical detection technique for monitoring antibody-antigen reactions at a quartz-liquid interface are described. The antibody is covalently immobilized on the optical surface of a planar, fused-quartz waveguide and reacted with antigen solution. A light beam is internally reflected within the waveguide and penetrates into the solution only a fraction of the wavelength of the incident light. This is the evanescent wave which interacts optically with the growing number of antigen-antibody complexes but minimally with the bulk solution. A two-site immunofluorescent assay for human IgG measurement is described using fluorescein as the label. The assay detection limit is approximately 0.8 micrograms/ml and individual fluorescence measurements are completed within 10 min. It is expected that this evanescent wave immunoassay should have wide applicability in both routine and research fields.