In magnetic nanoparticle (MNP)-clustering assays, a target molecule is bound to multiple receptors tethered onto MNPs, triggering MNP-clustering and leading to changes in the size of clusters. However, sandwich-type clustering requires multiple binding-sites on a target molecule, which is often unavailable for small-molecules. Furthermore, measuring magnetic properties as signals is not intrinsically selective regarding MNP-cluster size. Thus, the detection of few MNP-clusters is readily interfered by background signals from predominantly-existing single MNPs. Additionally, bulky and high-cost instruments limit the advancement of MNP-based assays. We report here a novel MNP-clustering small-molecule assay on an optical readout platform to overcome the limitations aforementioned with the following improvements. First, a facile MNP-clustering assay applicable to diverse small-molecules was realized by adopting an inhibition mechanism. Next, frequency-dependent optical measurements enabled us to resolve signals depending on the cluster size. Lastly, a low-cost and miniaturized optical readout setup was established by implementing a Blu-ray pickup head. Consequently, our low-cost optical biosensor using MNP-clustering facilitates high-resolution small-molecule assays. For experiments, aptamer-functionalized MNPs (Apt-MNPs) were first incubated with adenosine-5'-triphosphate (ATP) followed by adding MNPs with linker strands (linker-MNPs). The linker hybridizes with a region of aptamer sequences in the absence of ATP, forming MNP-clusters. Conversely, when aptamers are preoccupied by ATP inhibiting the hybridization, the cluster formation is hindered (Fig. 1). Consequently, higher ATP-concentrations result in smaller and fewer cluster formations. Blu-ray optical transmission measurements through an MNP-solution (Fig. 2) reveal that the 2nd harmonic component of the signal is related to the frequency of Brownian relaxation dynamics of MNPs induced by alternating magnetic field. Following measurement characterization (Fig. 3) and analyte-control experiments (Fig. 4), we demonstrated the ATP concentration dependent behavior of signals in micromolar ranges (Fig. 5). These results support that our MNP-clustering optical biosensor is capable of specific and quantitative detection of small-molecules.
Proceedings of the 14th Anniversary World Congress on Biosensors, 2014
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24th Anniversary World Congress on Biosensors, 2014