Abstract
Improving the signal-to-noise ratio (SNR) by amplifying the outputting signal or reducing nonspecific binding (NSB) are the key techniques in multiple immunoassay. Aiming at these issues, this paper presents an improved multiple indirect competitive immune surface-enhanced Raman scattering (ci-SERS) assay for the rapid screening of highly toxic rodenticides in food and biological samples, which ensured remarkable accuracy, ultra-sensitivity and reproducibility. The non-fouling polymer brush grafted magnetic beads (the MB@P-CyM) were prepared as multiple competitive recognition substrates after conjugating triplex haptens (the MB@P-CyM-hap). It was demonstrated that the particular 3D hair-like structures of P-CyM not only facilitate conjugate high-density hapten but reduce the steric hindrance from SERS probes recognition, thus enhancing SNB. On the other hand, Au nanoflowers (AuNFs) of high SERS activity were synthesized using a simple one-pot hydrazine reduction. For simultaneously detecting three highly toxic rodenticides, i.e., diphacinone (DPN), bromadiolone (BRD) and tetramine (TET), the obtained AuNFs were fabricated as a SERS-encoded nanoprobe cocktail after successively labeling mono-antibodies/Raman probes. By integrating the MB@P-CyM-hap with the SERS-encoded cocktail, a highly sensitive multiple SERS assay was achieved in less than 2 h with a limit of detection of 0.62 ng mL-1 for BRD, 0.42 ng mL-1 for TET and 1.37 ng mL-1 for DPN, respectively. The recoveries of these rodenticides in spiked food and biological samples were determined and ranged from 72 to 123%. Above all, the proposed modifications show remarkable improvements for high efficient multiple chemical toxin immunoassay.