Employment of Iron-Binding Protein from Haemophilus influenzae in Functional Nanopipettes for Iron Monitoring.

Because of the serious neurologic consequences of iron deficiency and iron excess in the brain, interest in the iron status of the central nervous system has increased significantly in the past decade. While iron plays an important role in many physiological processes, its accumulation may lead to diseases such as Huntington's, Parkinson's, and Alzheimer's. Therefore, it is important to develop methodologies that can monitor the presence of iron in a selective and sensitive manner. In this paper, we first showed the synthesis and characterization of the iron-binding protein (FBP) from Haemophilus influenzae, specific for ferrous ions. Subsequently, we employed this protein in our nanopipette platform and utilized it in functionalized nanoprobes to monitor the presence of ferrous ions. A suite of characterization techniques: absorbance spectroscopy, dynamic light scattering, and small-angle X-ray scattering were used for FBP. The functionalized Fe-nanoprobe calibrated in ferrous chloride enabled detection from 0.05 to 10 µM, and the specificity of the modified iron probe was evaluated by using various metal ion solutions.

Quantitative comparison of adsorption and desorption of commonly used sweeteners in the oral cavity.

Adsorption-desorption properties of different sweeteners in the oral cavity were evaluated using high performance liquid chromatography-based methodology. Three low calorie artificial sweeteners (aspartame, acesulfame potassium and sucralose), one steviol glycoside (rebaudioside A), and high fructose corn syrup (HFCS) were examined and compared with sucrose at pH 3 and 7 in a model beverage matrix. Results indicated that HFCS had the highest adsorption in the oral cavity, followed by rebaudioside A and the artificial sweeteners. The physicochemical interaction between sweeteners and salivary proteins did not affect the adsorption properties significantly as validated from a series of characterization techniques.

Nanopipettes as Monitoring Probes for the Single Living Cell: State of the Art and Future Directions in Molecular Biology.

Examining the behavior of a single cell within its natural environment is valuable for understanding both the biological processes that control the function of cells and how injury or disease lead to pathological change of their function. Single-cell analysis can reveal information regarding the causes of genetic changes, and it can contribute to studies on the molecular basis of cell transformation and proliferation. By contrast, whole tissue biopsies can only yield information on a statistical average of several processes occurring in a population of different cells. Electrowetting within a nanopipette provides a nanobiopsy platform for the extraction of cellular material from single living cells. Additionally, functionalized nanopipette sensing probes can differentiate analytes based on their size, shape or charge density, making the technology uniquely suited to sensing changes in single-cell dynamics. In this review, we highlight the potential of nanopipette technology as a non-destructive analytical tool to monitor single living cells, with particular attention to integration into applications in molecular biology.

Functionalized Quartz Nanopipette for Intracellular Superoxide Sensing: A Tool for Monitoring Reactive Oxygen Species Levels in Single Living Cell.

Reactive oxygen species (ROS), including superoxide radical anions, are vital components in numerous biological functions, including cell signaling and immune responses. Since ROS react with other biomolecules and oxidize them quickly, it is essential for cells to have superoxide-scavenging enzymes and other regulating enzymes that can catalyze the dismutation of superoxide radical anions into less damaging molecules. Otherwise, ROS overproduction can cause oxidative damage to DNA, proteins, cells, and tissues, damage that is associated with the pathogenesis of a range of neurodegenerative disorders, age-related diseases, and cancer. Understanding the relationship between superoxide and these disorders can help the development of innovative therapies for combating oxidative stress and degeneration of nerve cells. Although methods to quantify ROS already exist, they are indirect, destructive, ambiguous, and/or cannot provide real-time measurements in single cells. In this paper, we report a technique for sensing superoxide radical anions in single living cells using functionalized nanopipettes. These nanopipettes allow us to enter the cell as we measure intracellular ROS concentrations over time. We observed that these devices provide precise real-time measurements that are accurate and not possible to obtain with other conventional techniques.

DNA assay based on Nanoceria as Fluorescence Quenchers (NanoCeracQ DNA assay).

Functional nanomaterials with fluorescent or quenching abilities are important for the development of molecular probes for detection and studies of nucleic acids. Here, we describe a new class of molecular nanoprobes, the NanoCeracQ that uses nanoceria particles as a nanoquencher of fluorescent oligonucleotides for rapid and sensitive detection of DNA sequences and hybridization events. We show that nanoceria forms stable and reversible bionanoconjugates with oligonucleotides and can specifically recognize and detect DNA sequences in a single step. In absence of the target DNA, the nanoprobe produced minimal background fluorescence due to the high quenching efficiency of nanoceria. Competitive binding of the target induced a concentration dependent increase in the fluorescence signal due to hybridization and release of the fluorescent tag from the nanoparticle surface. The nanoprobe enabled sensitive detection of the complementary strand with a detection limit of 0.12 nM, using a single step procedure. The results show that biofunctionalized nanoceria can be used as a universal nanoquencher and nanosensing platform for fluorescent DNA detection and studies of nucleic acid interactions. This approach can find broad applications in molecular diagnostics, sensor development, gene expression profiling, imaging and forensic analysis.

Nano-Aptasensing in Mycotoxin Analysis: Recent Updates and Progress.

Recent years have witnessed an overwhelming integration of nanomaterials in the fabrication of biosensors. Nanomaterials have been incorporated with the objective to achieve better analytical figures of merit in terms of limit of detection, linear range, assays stability, low production cost, etc. Nanomaterials can act as immobilization support, signal amplifier, mediator and artificial enzyme label in the construction of aptasensors. We aim in this work to review the recent progress in mycotoxin analysis. This review emphasizes on the function of the different nanomaterials in aptasensors architecture. We subsequently relate their features to the analytical performance of the given aptasensor towards mycotoxins monitoring. In the same context, a critically analysis and level of success for each nano-aptasensing design will be discussed. Finally, current challenges in nano-aptasensing design for mycotoxin analysis will be highlighted.

ssDNA-Functionalized Nanoceria: A Redox-Active Aptaswitch for Biomolecular Recognition.

Quantification of biomolecular binding events is a critical step for the development of biorecognition assays for diagnostics and therapeutic applications. This paper reports the design of redox-active switches based on aptamer conjugated nanoceria for detection and quantification of biomolecular recognition. It is shown that the conformational transition state of the aptamer on nanoceria, combined with the redox properties of these particles can be used to create surface based structure switchable aptasensing platforms. Changes in the redox properties at the nanoceria surface upon binding of the ssDNA and its target analyte enables rapid and highly sensitive measurement of biomolecular interactions. This concept is demonstrated as a general applicable method to the colorimetric detection of DNA binding events. An example of a nanoceria aptaswitch for the colorimetric sensing of Ochratoxin A (OTA) and applicability to other targets is provided. The system can sensitively and selectivity detect as low as 0.15 × 10(-9) m OTA. This novel assay is simple in design and does not involve oligonucleotide labeling or elaborate nanoparticle modification steps. The proposed mechanism discovered here opens up a new way of designing optical sensing methods based on aptamer recognition. This approach can be broadly applicable to many bimolecular recognition processes and related applications.

Portable Nanoparticle-Based Sensors for Food Safety Assessment.

The use of nanotechnology-derived products in the development of sensors and analytical measurement methodologies has increased significantly over the past decade. Nano-based sensing approaches include the use of nanoparticles (NPs) and nanostructures to enhance sensitivity and selectivity, design new detection schemes, improve sample preparation and increase portability. This review summarizes recent advancements in the design and development of NP-based sensors for assessing food safety. The most common types of NPs used to fabricate sensors for detection of food contaminants are discussed. Selected examples of NP-based detection schemes with colorimetric and electrochemical detection are provided with focus on sensors for the detection of chemical and biological contaminants including pesticides, heavy metals, bacterial pathogens and natural toxins. Current trends in the development of low-cost portable NP-based technology for rapid assessment of food safety as well as challenges for practical implementation and future research directions are discussed.

Evaluation of the oxidase like activity of nanoceria and its application in colorimetric assays.

Nanomaterial-based enzyme mimics have attracted considerable interest in chemical analysis as alternative catalysts to natural enzymes. However, the conditions in which such particles can replace biological catalysts and their selectivity and reactivity profiles are not well defined. This work explored the oxidase like properties of nanoceria particles in the development of colorimetric assays for the detection of dopamine and catechol. Selectivity of the system with respect to several phenolic compounds, the effect of interferences and real sample analysis are discussed. The conditions of use such as buffer composition, selectivity, pH, reaction time and particle type are defined. Detection limits of 1.5 and 0.2µM were obtained with nanoceria for dopamine and catechol. The same assay could be used as a general sensing platform for the detection of other phenolics. However, the sensitivity of the method varies significantly with the particle type, buffer composition, pH and with the structure of the phenolic compound. The results demonstrate that nanoceria particles can be used for the development of cost effective and sensitive methods for the detection of these compounds. However, the selection of the particle system and experimental conditions is critical for achieving high sensitivity. Recommendations are provided on the selection of the particle system and reaction conditions to maximize the oxidase like activity of nanoceria.

A generic amplification strategy for electrochemical aptasensors using a non-enzymatic nanoceria tag.

We report a novel non-enzymatic nanocatalyst based approach to construct an electrochemical aptasensor involving the synergistic contribution of a nanoceria (nCe) tag and graphene oxide (GO). The aptamer was immobilized on the surface of a GO modified electrode. The target analyte was captured by the immobilized aptamer via a specific competitive mechanism between the free and the nCe labeled target. The electrochemical signal was generated by monitoring the electro-oxidation of a generic redox species upon reaction with the nCe tag. The signal was further amplified by the GO layer used as an electrode material to immobilize the aptamer and to increase the electron transfer at the electrode surface, further enhancing sensitivity of the assay. This strategy provides a universal platform for sensitive and specific detection of a wide spectrum of aptamer targets. Application of this new design for the electrochemical detection of Ochratoxin A (OTA) is demonstrated. Under optimal conditions, the aptasensor exhibited a linear response to OTA in the range 0.15-180 nM with a detection limit of 0.1 nM. The method has been successfully used for the detection of OTA in cereal samples. This design may offer a new methodology for sensitive and specific detection of a wide spectrum of analytes for medical, environmental and electronic applications.

Redox reactivity of cerium oxide nanoparticles against dopamine.

The interaction between dopamine and the redox active cerium oxide nanoparticles, or nanoceria was studied using a suite of spectroscopic and surface characterization methods. Changes in the chemical reactivity and concentration of dopamine upon exposure to nanoceria was assessed in aqueous solutions and a human physiological fluid--human serum. The results indicate strong attachment of dopamine to the nanoparticle surface through oxidation followed by chemisorption of the oxidative product with formation of a charge transfer complex. Such oxidation/surface adsorption processes between nanoceria and dopamine lead to a reduction of the concentration of free dopamine in aqueous environments. These findings suggest that the redox reactivity of nanoceria may alter dopamine levels in biological systems exposed to these particles and indicate the need for a comprehensive assessment of the potential neurological consequences that might result from intended or unintended exposure to these particles.