Blue silica nanoparticle-based colorimetric immunoassay for detection of Salmonella pullorum

Abstract

A colorimetric immunoassay based on blue silica nanoparticles (blue-SiNPs) was developed for quantitative detection of Salmonella pullorum (S. pullorum). In this method, blue-SiNPs were synthesized by doping C.I. reactive blue 21 into silica nanoparticles using a reverse microemulsion method. Blue-SiNPs functionalized with anti-S. pullorum were employed as detection probes. Magnetic nanoparticles (MNPs), employed as supports for the immobilization of polyclonal antibodies against S. pullorum, were used as capture probes. The sandwich structures of MNP-S. pullorum-blue-SiNPs were separated with a magnet and etched with NaOH. The C.I. reactive blue 21, released from silica nanoparticles, was used as a colorimetric indicator. The absorbance of C.I. reactive blue 21 at 675 nm is proportional to the concentration of S. pullorum. Under optimal conditions, the developed colorimetric immunoassay exhibited a wide dynamic range of 4.4 × 10² CFU mL⁻¹ to 4.4 × 10⁷ CFU mL⁻¹ toward S. pullorum with a detection limit of 4.4 × 10¹ CFU mL⁻¹. For application of the assay, this method is not influenced by the complex matrix of practical samples. The recoveries of S. pullorum from chicken liver samples were from 94.5% to 108% with a good correlation coefficient (R² = 0.9989) with those obtained by an official standard culture method. We also show that this colorimetric immunoassay can be carried out on a microplate reader with a 96-well plate. This method is particularly economic, simple, rapid, specific and has good stability. The new technology provided a basis for the detection of other pathogenic bacteria and viruses. Such a simple colorimetric immunoassay holds great potential as an on-site tool for clinical diagnosis of bacteria and viruses.