Applications of Hairpin DNA-Functionalized Gold Nanoparticles for Imaging mRNA in Living Cells

Molecular imaging agents are useful for imaging molecular processes in living systems in order to elucidate the function of molecular mediators in health and disease. Here, we demonstrate a technique for the synthesis, characterization, and application of hairpin DNA-functionalized gold nanoparticles (hAuNPs) as fluorescent hybridization probes for imaging mRNA expression and spatiotemporal dynamics in living cells. These imaging probes feature gold colloids linked to fluorophores via engineered oligonucleotides to resemble a molecular beacon in which the gold colloid serves as the fluorescence quencher in a fluorescence resonance energy transfer system. Target-specific hybridization of the hairpin oligonucleotide enables fluorescence de-quenching and subsequent emission with high signal to noise ratios. hAuNPs exhibit high specificity without adverse toxicity or the need for transfection reagents. Furthermore, tunability of hAuNP emission profiles by selection of spectrally distinct fluorophores enables multiplexed mRNA imaging applications. Therefore, hAuNPs are promising tools for imaging gene expression in living cells. As a representative application of this technology, we discuss the design and applications of hAuNP targeted against distinct matrix metalloproteinase enzymes for the multiplexed detection of mRNA expression in live breast cancer cells using flow cytometry and fluorescence microscopy.