Development of smart nanoparticle-aptamer sensing technology

Quantum dots (QDs) are excellent donors in Förster resonance energy transfer (FRET) based sensors because of their broad absorption and narrow symmetric emission. However, the strict requirement of a short donor-acceptor distance to achieve high FRET (hence sensitivity) has posed a significant challenge for QD-FRET-based sensors due to challenges associated with the preparation of QD conjugates that are both compact and highly stable. Consequently, most robust QD-FRET sensors are often too bulky to produce FRET efficiently, especially at low target-to-QD copy numbers. They have largely relied on increasing the target:QD ratio to achieve high FRET, making them undesirable and inefficient in situations of low target:QD copy numbers. Herein we report our work on the preparation of stable, compact and water-soluble QDs and their subsequent use in making compact, functional QD-DNA-based smart nanoparticle sensors for labelled and label-free DNA and protein detection. We have developed two strategies to prepare QD-DNA sensors: 1) via QD-thiolated DNA self-assembly, and 2) via covalent coupling between DNA and a QD surface ligand functional group. We found that thiolated DNA (fluorophore labelled) can self-assemble onto a 3-mercaptopropionic acid-capped QDs to produce highly efficient FRET (~80%) at a DNA:QD ratio of 1 : 1. However, this system suffers from strong non-specific adsorption and the self-assembled single-stranded (ss) DNA target is unable to hybridise to its complementary probe. More recently, we found that a dihydrolipoic acid-capped QD-ssDNA self-assembled system can hybridise to a labelled complementary probe to produce efficient FRET that can be exploited for labelled DNA probe quantification. Further, incorporating an anti-thrombin DNA aptamer to this system leads to a QD-DNA aptamer sensor that can specifically detect a 10 nM unlabelled protein probe (thrombin). The non-specific adsorption problem can be eliminated by introducing a poly(ethylene glycol) (PEG) linker to the QD capping ligands or by capping the QD with a chelating dendritic ligand. The resulting QD-DNA sensors can specifically detect 1 nM unlabelled or 35 pM labelled DNA probes using QD-sensitised dye FRET signals on a conventional fluorimeter. Extension of the DNA target to other functional DNAs or DNA/RNA aptamers should allow the development of a multi-functional QD-DNA platform suitable for biosensing, disease diagnosis and therapeutic applications.