Infrared optical immunosensor: application to the measurement of the herbicide atrazine

A new approach to optically transduce antigen-antibody association, needing no label, is described herein, taking advantage of the ability of reflection-absorption infrared (IR) spectroscopy to analyze organic thin films at the surface of reflective materials with high sensitivity. As a proof-of-principle, this new technique was applied to the immunodetection of the herbicide atrazine. Gold-coated chips were covered with a capture layer consisting of a protein derivative of the herbicide atrazine covalently bound to a self-assembled monolayer containing a carboxy-terminated thiolate. Successive binding of anti-atrazine antibody and secondary anti-rabbit immunoglobulin G antibody resulted in a change of the IR absorption properties of the organic film at the sensor surface. The two prominent amide I and II bands observed on the surface IR spectra were taken for semiquantitative analysis of the adsorbed protein amount. The presence of increasing amounts of atrazine resulted in the progressive inhibition of antibodies binding to the sensors, yielding a relative lower increase of the IR signals. The deduced standard curves displayed a sigmoidal shape typical of competitive inhibition assays. The test midpoint (IC(50)) and the limit of detection (IC(80)) were found to be in the nanomolar range and very close to those measured by an in-house enzyme-linked immunosorbent assay using the same antibody and the same antigen competitor.