Mohtashim H. Shamsi

Interactions of Metal Ions with DNA and Some Applications

Nucleic acids play a critical role in life as we know it. It contains the necessary information required for the structure and function of a living organisms. Metal ions play a critical role in stabilizing conformations. In the well-known double helix structure of DNA, metal ions stabilize a particular conformation that ensures storage and propagation of genetic information. Metal ions, however, can interact with various sites on nucleic acids. Moreover, metal coordination can have a tremendous impact on the structure, conformation, stability and the electronic properties of the nucleic acids. The interactions are controlled by the relative affinity of metal ion coordination to the negatively charged phosphodiester backbone versus binding to other donor sites located in the nucleobases. The canonical Watson–Crick base pairs (A-T and G-C) as well as non-canonical base pairs (Hoogsteen and wobble) and mismatched pairs are often sites for metal ion interactions. In this review, an overview will be provided of the structure of different forms of nucleic acids (DNA and RNA) and the impact of different metal ions on their stability and structure. In addition, the recent applications of metal-DNA interactions in nanotechnology, biosensor and bioelectronics will also be discussed along with some therapeutic applications of metal complexes.

Electrochemiluminescence on digital microfluidics for microRNA analysis.

Electrochemiluminescence (ECL) is a sensitive analytical technique with great promise for biological applications, especially when combined with microfluidics. Here, we report the first integration of ECL with digital microfluidics (DMF). ECL detectors were fabricated into the ITO-coated top plates of DMF devices, allowing for the generation of light from electrically excited luminophores in sample droplets. The new system was characterized by making electrochemical and ECL measurements of soluble mixtures of tris(phenanthroline)ruthenium(II) and tripropylamine (TPA) solutions. The system was then validated by application to an oligonucleotide hybridization assay, using magnetic particles bearing 21-mer, deoxyribose analogues of the complement to microRNA-143 (miRNA-143). The system detects single nucleotide mismatches with high specificity, and has a limit of detection of 1.5 femtomoles. The system is capable of detecting miRNA-143 in cancer cell lysates, allowing for the discrimination between the MCF-7 (less aggressive) and MDA-MB-231 (more aggressive) cell lines. We propose that DMF-ECL represents a valuable new tool in the microfluidics toolbox for a wide variety of applications.

Investigation of the Utility of Complementary Electrochemical Detection Techniques to Examine the in Vitro Affinity of Bacterial Flagellins for a Toll-Like Receptor 5 Biosensor

An initial investigation of the fabrication of a novel biosensor utilizing toll-like receptor 5 (TLR5) has been conducted. The detection assay using this sensor platform has been carried out using two complementary electrochemical techniques. The electrochemical properties of the modified bare gold surface following TLR5 immobilization were characterized. The electrochemical response to changes in the sensor film resistance and electron charge-transfer permittivity triggered by independent exposures to flagellins from Salmonella typhimurium (S. typhimurium) and Bacillus subtilis (B. subtilis) were examined and observed. The quantified film resistance data gathered using electrochemical impedance spectroscopy (EIS) over a macroscopic scale are in significant agreement with the corresponding electron charge-transfer permittivity measured locally by scanning electrochemical microscopy (SECM). Unlike other sensors that exploit pathogen recognition elements, TLR5 biosensors have the potential to carry out broad-spectrum detection of flagellated bacterial pathogens in near real time. This broad-spectrum detection platform is a significant step toward the development of fast, inexpensive clinical tools for early warning diagnoses and immediate on-site treatment.

Integrated digital microfluidic platform for voltammetric analysis.

Digital microfluidics (DMF) is an emerging technique for manipulating small volumes of liquids. DMF is particularly well suited for analytical applications as it allows automated handling of discrete samples, and it has been integrated with several inline analysis techniques. However, examples of the integration of DMF with electroanalytical methods are notably scarce, and those that have been reported rely on external electrodes that impose limitations on complexity. To combine the full capabilities of DMF with voltammetry, we designed a platform featuring a three-electrode electrochemical cell integrated in a microfabricated DMF device, removing the need for external electrodes and allowing for complete droplet control. The performance of the DMF/voltammetry system is comparable to that of a commercial screen printed electrode, and the new platform was applied to generating a calibration series for acetaminophen with a limit of detection of 76 μM and good precision (4% average RSD), all with minimal human intervention. We propose that this platform and variations thereof may be a useful new tool for microscale electroanalysis and will be a complementary system to existing inline analysis techniques for DMF.

DNA Films Containing the Artificial Nucleobase Imidazole Mediate Charge Transfer in a Silver(I)-Responsive Way

The first sequence-dependent study of DNA films containing metal-mediated base pairs was performed to investigate the charge transfer resistance (RCT ) of metal-modified DNA. The imidazole (Im) deoxyribonucleoside was chosen as a highly AgI -specific ligandoside for the formation of Im-AgI -Im complexes within the duplexes. This new class of site-specifically metal-modified DNA films was characterized by UV, circular dichroism (CD), and X-ray photoelectron spectroscopy (XPS). The electrochemical properties of these systems were investigated by means of electron impedance spectroscopy and scanning electrochemical microscopy. Taken together, these experiments indicated that the incorporation of AgI ions into the DNA films leads to reduced electron transfer through the DNA films. A simple device was proposed that can be switched reversibly between two distinct states with different charge transfer resistance.

Wax patterned microwells for stem cell fate study

The fabrication of cost effective paper-based analytical devices by wax printing has recently become popular, by and large, using cellulose filter papers. Paper-based devices need higher temperature to form hydrophobic barrier across paper substrate, rely on large working channels (≥500 μm) for liquid handling, and exhibit lower efficiency (∼50%) of sample mobility. Such limitations confine applications of wax based fabrication. In this work, we report printability, fidelity, and application of wax micropatterns on a non-cellulosic, non-fibrous, and non-porous polyethylene terephthalate based substrate (mPET). Resolution of wax printing on mPET was found to be 120 μm for line and 60 μm for channel micropatterns. The wax micropatterns can sustain heat and retain their structural integrity at melting temperature of wax and above (≥120 °C). In application, wax microwells were patterned on the new substrate in a high throughput fashion, which formed a suitable niche for mouse embryonic stem cell (mESC) culture either to maintain self-renewal or direct differentiation. This study will open a new direction in wax printing applications not only as a low-cost but a multipurpose fabrication tool.

Scanning Electrochemical Microscopy: A Multiplexing Tool for Electrochemical DNA Biosensing

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