Abstract
A turn-on competitive immunoassay using a low-cost Blu-ray optomagnetic setup and two differently sized magnetic particles (micron-sized particles acting as capture particles and nano-sized particles acting as detection particles) is here presented. For Salmonella detection, a limit of detection of 8×10(4)CFU/mL is achieved within a total assay time of 3h. The combination of a competitive strategy and an optomagnetic setup not only enables a turn-on read-out format, but also results in a sensitivity limit about a factor of 20 times lower than of volumetric magnetic stray field detection device based immunoassays. The improvement of sensitivity is enabled by the formation of immuno-magnetic aggregates providing steric hindrance protecting the interior binding sites from interaction with the magnetic nanoparticle labels. The formation of immuno-magnetic aggregates is confirmed by fluorescence microscopy. The system exhibits no visible cross-reaction with other common pathogenic bacteria, even at concentrations as high as 10(7)CFU/mL. Furthermore, we present results when using the setup for a qualitative and homogeneous biplex immunoassay of Escherichia coli and Salmonella typhimurium.
Highlights
A turn-on competitive immunoassay using a low-cost Blu-ray optomagnetic setup and two differently sized magnetic particles is here presented
E. coli directly binds to Ab250 nm detection magnetic nanoparticles (MNPs), i.e. a direct detection strategy, while simultaneously S. typhimurium binds to the antibody coated μm magnetic particles (Ab-MPs) to form immuno-magnetic aggregates, i.e. a competitive strategy
If S. typhimurium cells and/or fragments are suspended in the sample, these entities will react with the biotinylated antibodies on the surface of the MPs and sterically prevent these antibodies from interacting with the subsequently added detection nanoparticles
Summary
Where sophisticated lab facilities are not available (Kelley et al, 2014; Brandao et al, 2015). On the other hand, when employing a clinical affinity capture strategy, e.g. a bacterial immunoassay, the sensitivity of those devices (LOD of higher than 106 CFU/mL) is inferior compared to that of a miniaturized nuclear magnetic resonance (NMR) system (LOD of 103 CFU/mL), which benefits from a built-in signal amplification (since millions of water molecules can be affected by one individual magnetic particle) (Grossman et al, 2004; Chen et al, 2015). All those aforementioned biosensors require expensive instrumentation. It is worth mentioning that the cost of this read-out system is very low in comparison to other devices, e.g. only a few percent of the cost for a miniaturized NMR device (about 50,000€)
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