A Comprehensive Screen of Metal Oxide Nanoparticles for DNA Adsorption, Fluorescence Quenching, and Anion Discrimination

This document is the Accepted Manuscript version of a Published Work that appeared in final form in Applied Materials & Interfaces, copyright © American Chemical Society after peer review and technical editing by publisher. To access the final edited and published work see http://dx.doi.org/10.1021/acsami.5b08004Although DNA has been quite successful in metal cation detection, anion detectioin remains challenging because of the charge repulsion. Metal oxides represent a very important class of materials, and different oxides might interact with anions differently. In this work, a comprehensive screen of common metal oxide nanoparticles (MONPs) was carried out for their ability to adsorb DNA, quench fluorescence, and release adsorbed DNA in the presence of target anions. A total of 19 MONPs were studied, including Al2O3, CeO2, CoO, Co3O4, Cr2O3, Fe2O3, Fe3O4, In2O3, ITO, Mn2O3, NiO, SiO2, SnO2, a-TiO2 (anatase), r-TiO2 (rutile), WO3, Y2O3, ZnO, ZrO2. These MONPs have different DNA adsorption affinity. Some adsorb DNA without quenching the fluorescence, while others strongly quench adsorbed fluorophores. They also display different affinity toward anions probed by DNA desorption. Finally, CeO2, Fe3O4, and ZnO were used to form a sensor array to discriminate phosphate, arsenate, and arsenite from the rest using linear discriminant analysis. This study not only provides a solution for anion discrimination using DNA as a signaling molecule but also provides insights into the interface of metal oxides and DNA.Natural Sciences and Engineering Research Council || Discovery and Strategic Project Grant: STPGP-447472-2013 05576

Accelerating peroxidase mimicking nanozymes using DNA

DNA-capped iron oxide nanoparticles are nearly 10-fold more active as a peroxidase mimic for TMB oxidation than naked nanoparticles. To understand the mechanism, the effect of DNA length and sequence is systematically studied, and other types of polymers are also compared. This rate enhancement is more obvious with longer DNA and, in particular, poly-cytosine. Among the various polymer coatings tested, DNA offers the highest rate enhancement. A similar acceleration is also observed for nanoceria. On the other hand, when the positively charged TMB substrate is replaced by the negatively charged ABTS, DNA inhibits oxidation. Therefore, the negatively charged phosphate backbone and bases of DNA can increase TMB binding by the iron oxide nanoparticles, thus facilitating the oxidation reaction in the presence of hydrogen peroxide.University of Waterloo || Canadian Foundation for Innovation || Natural Sciences and Engineering Research Council || Ontario Ministry of Research and Innovation |

DNA Adsorption by Indium Tin Oxide (ITO) Nanoparticles

This document is the Accepted Manuscript version of a Published Work that appeared in final form in Langmuir, copyright © American Chemical Society after peer review and technical editing by publisher. To access the final edited and published work see http://dx.doi.org/10.1021/la503917jThe high conductivity and optical transparency of indium tin oxide (ITO) has made it a popular material in the electronic industry. Recently, its application in biosensors is also explored. To understand its biointerface chemistry, we herein investigate its interaction with fluorescently labeled single-stranded oligonucleotides using ITO nanoparticles (NPs). The fluorescence of DNA is efficiently quenched after adsorption, and the interaction between DNA and ITO NPs is strongly dependent on the surface charge of ITO. At low pH, the ITO surface is positively charged to afford a high DNA adsorption capacity. Adsorption is also influenced by the sequence and length of DNA. For its components, In2O3 adsorbs DNA more strongly while SnO2 repels DNA at neutral pH. The DNA adsorption property of ITO is an averaging result from both components. DNA adsorption is confirmed to be mainly by the phosphate backbone via displacement experiments using free phosphate or DNA bases. Last, DNA-induced DNA desorption by forming duplex DNA is demonstrated on ITO, while the same reaction is more difficult to achieve on other metal oxides including CeO2, TiO2, and Fe3O4 because these particles adsorb DNA more tightly.University of Waterloo || Canadian Foundation for Innovation || Natural Sciences and Engineering Research Council || Ontario Ministry of Research and Innovation |

DNA adsorption by magnetic iron oxide nanoparticles and its application for arsenate detection

Iron oxide nanoparticles adsorb fluorescently labeled DNA oligonucleotides via the backbone phosphate and quench fluorescence. Arsenate displaces adsorbed DNA to increase fluorescence, allowing detection of arsenate down to 300 nM. This is a new way of using DNA: analyte recognition relies on its phosphate instead of the bases

Fluorescent sensors using DNA-functionalized graphene oxide

The final publication is available at Springer via http://dx.doi.org/10.1007/s00216-014-7888-3In the past few years, graphene oxide (GO) has emerged as a unique platform for developing DNA-based biosensors, given the DNA adsorption and fluorescence-quenching properties of GO. Adsorbed DNA probes can be desorbed from the GO surface in the presence of target analytes, producing a fluorescence signal. In addition to this initial design, many other strategies have been reported, including the use of aptamers, molecular beacons, and DNAzymes as probes, label-free detection, utilization of the intrinsic fluorescence of GO, and the application of covalently linked DNA probes. The potential applications of DNA-functionalized GO range from environmental monitoring and cell imaging to biomedical diagnosis. In this review, we first summarize the fundamental surface interactions between DNA and GO and the related fluorescence-quenching mechanism. Following that, the various sensor design strategies are critically compared. Problems that must be overcome before this technology can reach its full potential are described, and a few future directions are also discussed.University of Waterloo || Natural Sciences and Engineering Research Council || Ontario Ministry of Research and Innovation || Foundation for Shenghua Scholar || National Natural Science Foundation of China || Grant No. 81301258, 21301195 Postdoctoral Science Foundation of Central South University and Hunan province ||Grant No. 124896 China Postdoctoral Science Foundation || Grant No. 2013M540644 Hunan Provincial Natural Science Foundation of China || Grant No. 13JJ4029 Specialized Research Fund for the Doctoral Program of Higher Education of China || Grant No. 2013016212007

Cation-Size-Dependent DNA Adsorption Kinetics and Packing Density on Gold Nanoparticles: An Opposite Trend

This document is the Accepted Manuscript version of a Published Work that appeared in final form in Langmuir, copyright © American Chemical Society after peer review and technical editing by publisher. To access the final edited and published work see Liu, B., Kelly, E. Y., & Liu, J. (2014). Cation-Size-Dependent DNA Adsorption Kinetics and Packing Density on Gold Nanoparticles: An Opposite Trend. Langmuir, 30(44), 13228–13234. https://doi.org/10.1021/la503188hThe property of DNA is strongly influenced by counterions. Packing a dense layer of DNA onto a gold nanoparticle (AuNP) generates an interesting colloidal system with many novel physical properties such as a sharp melting transition, protection of DNA against nucleases, and enhanced complementary DNA binding affinity. In this work, the effect of monovalent cation size is studied. First, for free AuNPs without DNA, larger group 1A cations are more efficient in inducing their aggregation. The same trend is observed with group 2A metals using AuNPs capped by various self-assembled monolayers. After establishing the salt range to maintain AuNP stability, the DNA adsorption kinetics is also found to be faster with the larger Cs+ compared to the smaller Li+. This is attributed to the easier dehydration of Cs+, and dehydrated Cs+ might condense on the AuNP surface to reduce the electrostatic repulsion effectively. However, after a long incubation time with a high salt concentration, Li+ allows ∼30% more DNA packing compared to Cs+. Therefore, Li+ is more effective in reducing the charge repulsion among DNA, and Cs+ is more effective in screening the AuNP surface charge. This work suggests that physicochemical information at the bio/nanointerface can be obtained by using counterions as probes.University of Waterloo || Canadian Foundation for Innovation || Natural Sciences and Engineering Research Council || Ontario Ministry of Research and Innovation |

Characterization of glucose oxidation by gold nanoparticles using nanoceria

The final publication is available at Elsevier via http://dx.doi.org/10.1016/j.jcis.2014.04.025." © 2014. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/Gold nanoparticles (AuNPs) can oxidize glucose, producing hydrogen peroxide and gluconic acid, which are the same products as those generated by glucose oxidase (GOx). In this regard, AuNPs are a nanozyme. Herein, a new colorimetric method is developed to understand the surface chemistry of gold nanoparticles for this oxidation reaction. The color of nanoceria is changed to yellow by the hydrogen peroxide generated during glucose oxidation. Using this assay, we find that adsorption of small molecules such as citrate does not deactivate AuNPs, while adsorption of polymers including serum proteins and high molecular weight polyethylene glycol inhibits glucose oxidation. In addition to glucose, AuNPs can also oxidize galactose. Therefore, this reaction is unlikely to be directly useful for glucose detection for biomedical applications. On the other hand, AuNPs might serve as a general oxidase for a broad range of substrates. The glucose oxidation reaction is slower at lower pH. Since the reaction generates an acid product, glucose oxidation becomes slower as the reaction proceeds. The effects of temperature, AuNP size, and reaction kinetics have been systematically studied. This work provides new insights regarding the surface chemistry of AuNPs as a nanozyme.University of Waterloo || Canadian Foundation for Innovation || Natural Sciences and Engineering Research Council || Ontario Ministry of Research and Innovation |

Mechanisms of DNA Sensing on Graphene Oxide

This document is the Accepted Manuscript version of a Published Work that appeared in final form in Analytical Chemistry copyright © American Chemical Society after peer review and technical editing by publisher. To access the final edited and published work see Liu, B., Sun, Z., Zhang, X., & Liu, J. (2013). Mechanisms of DNA Sensing on Graphene Oxide. Analytical Chemistry, 85(16), 7987–7993. https://doi.org/10.1021/ac401845pAdsorption of a fluorophore-labeled DNA probe by graphene oxide (GO) produces a sensor that gives fluorescence enhancement in the presence of its complementary DNA (cDNA). While many important analytical applications have been demonstrated, it remains unclear how DNA hybridization takes place in the presence of GO, hindering further rational improvement of sensor design. For the first time, we report a set of experimental evidence to reveal a new mechanism involving nonspecific probe displacement followed by hybridization in the solution phase. In addition, we show quantitatively that only a small portion of the added cDNA molecules undergo hybridization while most are adsorbed by GO to play the displacement role. Therefore, it is possible to improve signaling by raising the hybridization efficiency. A key innovation herein is using probes and cDNA with a significant difference in their adsorption energy by GO. This study offers important mechanistic insights into the GO/DNA system. At the same time, it provides simple experimental methods to study the biomolecular reaction dynamics and mechanism on a surface, which may be applied for many other biosensor systems.University of Waterloo || Canadian Foundation for Innovation || Natural Sciences and Engineering Research Council || Ontario Ministry of Research and Innovation |

DNA/Metal Oxide Nanoconjugates: Fundamental Understandings and Analytical Applications

DNA-functionalized nanomaterials have shown versatile applications in biosensor development, biomedical diagnostics, therapy, and catalysis. DNA is attractive for this purpose for its programmable structure, molecular recognition function, and ease of modification. Various nanomaterials, including noble metals, carbons, metal oxides, soft polymeric nanostructures, and metal organic frameworks have been conjugated with DNA. Among them, metal oxide nanoparticles (MONPs) exhibit unique magnetic, catalytic, and surface properties. Most previously reported DNA/MONP conjugates were prepared with the help of surface coating layers or linkers. While such conjugation provides stable hybrid materials, the intrinsic surface properties of MONPs are often masked. The primary focus of this thesis is to interface DNA oligonucleotides with pristine MONPs to provide critical insights into the fundamental understandings at these bio-nano interfaces and to design functional biosensors towards environmentally and biologically important analytes. In Chapter 2 the interaction between indium-doped tin oxide nanoparticles (ITO NPs) and fluorescently labeled single-stranded DNA (ssDNA) is systematically studied. While electrochemical and photochemical biosensors based on ITO for DNA detection have been developed, little is known about the biointerface chemistry. The DNA adsorption and fluorescence quenching capability of ITO NPs is first confirmed. Salt concentration, pH, DNA sequence and length affect DNA adsorption. The adsorption mechanism is found to be through the phosphate backbone using displacement assays. ITO NPs but not In2O3 can discriminate ssDNA and double stranded DNA (dsDNA) based on the difference in their chain flexibility. In Chapter 3, the interaction between fluorescently labeled DNA and iron oxide nanoparticles is investigated. Fe3O4 NPs adsorb DNA via the phosphate backbone and quench the fluorescence. With the strong affinity between arsenate and Fe3O4, a highly sensitive arsenate sensor is demonstrated based on the displacement of fluorescently labeled DNA by arsenate. Arsenate displaces adsorbed DNA to increase fluorescence, allowing the detection of arsenate down to 300 nM. The sensor design represents a new way of using DNA: analyte recognition relying on metal oxide while DNA is used only as a signaling molecule. In Chapter 4, following the work in Chapter 2 and 3, a total of 19 MONPs are screened for their ability to adsorb DNA, quench fluorescence, and release adsorbed DNA in the presence of a few common anions. These MONPs have different fluorescence quenching properties, DNA adsorption affinity, and different sensitivity toward anions probed by DNA desorption. Finally, CeO2, Fe3O4, and ZnO are used to form a sensor array to discriminate phosphate, arsenate, and arsenite from the rest using the linear discriminant analysis method. The study not only provides a solution for anion discrimination using MONPs and DNA but also insights into the interface of metal oxides and DNA. In Chapter 5, a fluorescently labeled DNA is used as a probe to investigate the interaction between a biologically important molecule, H2O2, and a nanozyme, nanoceria. Nanoceria has been previously reported to bind DNA strongly. I demonstrate that the adsorbed DNA can be readily displaced by H2O2, resulting in over 20-fold fluorescence enhancement. The displacement mechanism instead of oxidative DNA cleavage is confirmed by denaturing gel electrophoresis and surface group pKa measurements. This system can sensitively detect H2O2 down to 130 nM. When coupled with glucose oxidase, glucose is detected down to 8.9 µM in buffer. Detection in serum is also achieved with results comparable with that from a commercial glucose meter. With an understanding of the ligand role of H2O2, new applications in rational materials design, sensor development, and drug delivery can be further exploited. In Chapter 6, I demonstrate the feasibility of using DNA in promoting the peroxidase activity of iron oxide nanoparticles. The effect of DNA length, sequence, surface coating are systematically studied. The rate enhancement is more significant with longer DNA. The negatively charged phosphate backbone and bases of DNA can increase the substrate binding, thus facilitating the oxidation reaction in the presence of H2O2. The role of DNA in modulating the peroxidase activity of iron oxide provides insights into the mechanism the nanozymes. Overall, the adsorption mechanism of DNA by various oxides, the controlling of the catalytic activity of oxides, and the related biosensor applications have been extensively studied in this thesis

Dissecting Colloidal Stabilization Factors in Crowded Polymer Solutions by Forming Self-Assembled Monolayers on Gold Nanoparticles

This document is the Accepted Manuscript version of a Published Work that appeared in final form in Langmuir, copyright © American Chemical Society after peer review and technical editing by publisher. To access the final edited and published work see [insert ACS Articles on Request author-directed link to Published Work, see Lang, N. J., Liu, B., Zhang, X., & Liu, J. (2013). Dissecting Colloidal Stabilization Factors in Crowded Polymer Solutions by Forming Self-Assembled Monolayers on Gold Nanoparticles. Langmuir, 29(20), 6018–6024. https://doi.org/10.1021/la3051093An ideal colloidal system should be highly stable in a diverse range of buffer conditions while still retaining its surface accessibility. We recently reported that dispersing citrate-capped gold nanoparticles (AuNPs) in polymers, such as polyethylene glycol (PEG), can achieve such a goal because of contributions from depletion stabilization. Because AuNPs can weakly adsorb PEG to exert steric stabilization and the remaining citrate can impart charge stabilization, the extent of the contribution of depletion stabilization is unclear. In this work, we aim to dissect the contribution of each stabilizing factor. This is achieved by coating AuNPs with a layer of thiolated compound, which inhibits the adsorption of PEG and also allows for the control of surface charge. We found that depletion stabilization alone was insufficient to stabilize AuNPs at room temperature. However, when working together with other stabilization mechanisms, ultrahigh stability can be achieved. The size of both AuNPs and PEG was systematically varied, and the trends were compared to theoretical calculations. Finally, we report the importance of the surface chemistry of commercial AuNPs.University of Waterloo || Canadian Foundation for Innovation || Natural Sciences and Engineering Research Council || Ontario Ministry of Research and Innovation |