Magnetic bead-based fluorescence immunoassay for aflatoxin B1 in food using biofunctionalized rhodamine B-doped silicananoparticles

Abstract

A simple and sensitive fluorescence immunoassay for the detection of aflatoxin B₁ (AFB₁, as a model compound) in food was developed using AFB₁–bovine serum albumin conjugate (AFB₁–BSA)-functionalized magnetic beads as immunosensing probes. The recognition elements were prepared by doping of rhodamine B (RB) fluorophore into silica nanoparticles followed by immobilization of monoclonal anti-AFB₁ antibodies on the silica shell. Based on a competitive-type immunoassay format, the assay was performed both in low-binding polypropylene 96-well microtiter plates (MTPs) and in an automated sequential injection (SI) format. Similar detection limit (LOD) of 0.2 ng mL⁻¹vs. 0.1 ng mL⁻¹ but narrower dynamic working linear range of 0.5–7 ng mL⁻¹vs. 0.5–30 ng mL⁻¹ was obtained toward AFB₁ standards with the flow setup compared to the MTP format. Intra-batch assay precision was substantially improved (≤5.3% vs. ≤8.7%) by resorting to the SI manifold. The proposed method features unbiased identification of negative (blank) and positive samples. No significant differences at the 95% confidence level were encountered in the analysis of naturally contaminated peanut samples between the proposed immunoassay and liquid chromatography for determination of AFB₁.