title: Applications of Hairpin DNA-Functionalized Gold Nanoparticles for Imaging mRNA in Living Cells.
abstract: 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.
authors.affiliation: Vanderbilt University, Nashville, TN, United States.
authors.forename: S R
authors.initials: SR
authors.lastname: Jackson
authors.affiliation: Vanderbilt University, Nashville, TN, United States.
authors.forename: A C
authors.initials: AC
authors.lastname: Wong
authors.affiliation: Vanderbilt University, Nashville, TN, United States.
authors.forename: A R
authors.initials: AR
authors.lastname: Travis
authors.affiliation: Hunter College, City University of New York, New York, NY, United States.
authors.forename: I E
authors.initials: IE
authors.lastname: Catrina
authors.affiliation: Hunter College, City University of New York, New York, NY, United States; Program in Molecular, Cellular, and Developmental Biology, and Program in Biochemistry, The Graduate Center, City University of New York, New York, NY, United States.
authors.forename: D P
authors.initials: DP
authors.lastname: Bratu
authors.affiliation: Vanderbilt University, Nashville, TN, United States.
authors.forename: D W
authors.initials: DW
authors.lastname: Wright
authors.affiliation: Pharma Research and Early Development (pRED), F. Hoffman-La Roche Ltd., Basel, Switzerland. Electronic address: ash.jayagopal@gmail.com.
authors.forename: A
authors.initials: A
authors.lastname: Jayagopal
mesh_terms: D001483:Base Sequence; D045744:Cell Line, Tumor; D002470:Cell Survival; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D005434:Flow Cytometry / Q000379:methods; D005456:Fluorescent Dyes / Q000737:chemistry*; D006046:Gold / Q000737:chemistry*; D006801:Humans; D018613:Microscopy, Confocal / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods; D053758:Nanoparticles / Q000737:chemistry*; D009693:Nucleic Acid Hybridization / Q000379:methods; D015345:Oligonucleotide Probes / Q000737:chemistry* / Q000235:genetics; D061848:Optical Imaging / Q000379:methods; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics
chemical_list: D005456:Fluorescent Dyes; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D006046:Gold; D004247:DNA
keywords: DNA oligonucleotides; FRET; Fluorescence imaging; Fluorescent probes; Gene expression; Live cell imaging; Molecular beacons; Nanoparticles; hAuNP; mRNA