Competitive colorimetric triazophos immunoassay employing magnetic microspheres and multi-labeled gold nanoparticles along with enzymatic signal enhancement

Abstract

The authors have developed a competitive immunoassay for the model pesticide triazophos (TRIAZ). The method is based on the use of only one monoclonal antibody immobilized on multi-labeled gold nanoparticle (AuNP) probes. The immunoassay uses two sets of probe. The first is a multi-labeled AuNP probe that carries monoclonal antibody (mcAb), single-stranded DNA (ssDNA) and horse radish peroxidase (HRP). The second is a magnetic microparticle (MMP) probe that is obtained by coating MMPs with ovalbumin coupled to TRIAZ. Free TRIAZ and MMP-immobilized TRIAZ compete for binding to the mcAb on the surface of the AuNPs. Because TRIAZ is a rather small molecule, it cannot be bound by two antibodies. The competitive immunoassay overcomes the limitations of small molecule detection using one kind of mcAb only. Sensitive transduction of the immunoreaction is accomplished by enzymatically catalyzed amplified. TRIAZ was quantified by a conventional ELISA and by the immunoassay presented here. Both method are highly sensitive and well reproducible. The new assay has a linear response in the 15 ng L−1 to 40 μg L−1 TRIAZ concentration range, and a 14 ng L−1 limit of detection which make it more sensitive than the ELISA. The recovery rate in case of spiked samples ranges from 78.4 to 105%, and the RSD is <20%. A good correlation was further established between the results of the immunoassay and those of GC-MS analysis.