Aeraj ul Haque

Electrochemical Biosensor of Nanocube- Augmented Carbon Nanotube Networks

Networks of single-walled carbon nanotubes (SWCNTs) decorated with Au-coated Pd (Au/Pd) nanocubes are employed as electrochemical biosensors that exhibit excellent sensitivity (2.6 mA mM(-1) cm(-2)) and a low estimated detection limit (2.3 nM) at a signal-to-noise ratio of 3 (S/N = 3) in the amperometric sensing of hydrogen peroxide. Biofunctionalization of the Au/Pd nanocube-SWCNT biosensor is demonstrated with the selective immobilization of fluorescently labeled streptavidin on the nanocube surfaces via thiol linking. Similarly, glucose oxidase (GOx) is linked to the surface of the nanocubes for amperometric glucose sensing. The exhibited glucose detection limit of 1.3 muM (S/N = 3) and linear range spanning from 10 muM to 50 mM substantially surpass similar CNT-based biosensors. These results, combined with the structures compatibility with a wide range of biofunctionalization procedures, would make the nanocube-SWCNT biosensor exceptionally useful for glucose detection in diabetic patients and well suited for a wide range of amperometric detection schemes for clinically important biomarkers.

A comparative study of enzyme immobilization strategies for multi-walled carbon nanotube glucose biosensors.

This work addresses the comparison of different strategies for improving biosensor performance using nanomaterials. Glucose biosensors based on commonly applied enzyme immobilization approaches, including sol-gel encapsulation approaches and glutaraldehyde cross-linking strategies, were studied in the presence and absence of multi-walled carbon nanotubes (MWNTs). Although direct comparison of design parameters such as linear range and sensitivity is intuitive, this comparison alone is not an accurate indicator of biosensor efficacy, due to the wide range of electrodes and nanomaterials available for use in current biosensor designs. We proposed a comparative protocol which considers both the active area available for transduction following nanomaterial deposition and the sensitivity. Based on the protocol, when no nanomaterials were involved, TEOS/GOx biosensors exhibited the highest efficacy, followed by BSA/GA/GOx and TMOS/GOx biosensors. A novel biosensor containing carboxylated MWNTs modified with glucose oxidase and an overlying TMOS layer demonstrated optimum efficacy in terms of enhanced current density (18.3 ± 0.5 µA mM(-1) cm(-2)), linear range (0.0037-12 mM), detection limit (3.7 µM), coefficient of variation (2%), response time (less than 8 s), and stability/selectivity/reproducibility. H(2)O(2) response tests demonstrated that the most possible reason for the performance enhancement was an increased enzyme loading. This design is an excellent platform for versatile biosensing applications.

Electrochemical Glucose Biosensor of Platinum Nanospheres Connected by Carbon Nanotubes

Background: Glucose biosensors comprised of nanomaterials such as carbon nanotubes (CNTs) and metallic nanoparticles offer enhanced electrochemical performance that produces highly sensitive glucose sensing. This article presents a facile biosensor fabrication and biofunctionalization procedure that utilizes CNTs electrochemically decorated with platinum (Pt) nanospheres to sense glucose amperometrically with high sensitivity. Method: Carbon nanotubes are grown in situ by microwave plasma chemical vapor deposition (MPCVD) and electrochemically decorated with Pt nanospheres to form a CNT/Pt nanosphere composite biosensor. Carbon nanotube electrodes are immobilized with fluorescently labeled bovine serum albumin (BSA) and analyzed with fluorescence microscopy to demonstrate their biocompatibility. The enzyme glucose oxidase (GOX) is immobilized onto the CNT/Pt nanosphere biosensor by a simple drop-coat method for amperometric glucose sensing. Results: Fluorescence microscopy demonstrates the biofunctionalization capability of the sensor by portraying adsorption of fluorescently labeled BSA unto MPCVD-grown CNT electrodes. The subsequent GOX-CNT/Pt nanosphere biosensor demonstrates a high sensitivity toward H2O2 (7.4 μA/mM/cm2) and glucose (70 μA/mM/cm2), with a glucose detection limit and response time of 380 nM (signal-to-noise ratio = 3) and 8 s (t90%), respectively. The apparent Michaelis-Menten constant (0.64 mM) of the biosensor also reflects the improved sensitivity of the immobilized GOX/nanomaterial complexes. Conclusions: The GOX-CNT/Pt nanosphere biosensor outperforms similar CNT, metallic nanoparticle, and more conventional carbon-based biosensors in terms of glucose sensitivity and detection limit. The biosensor fabrication and biofunctionalization scheme can easily be scaled and adapted for microsensors for physiological research applications that require highly sensitive glucose sensing.

RNA-seq analysis identifies potential modulators of gravity response in spores of Ceratopteris (Parkeriaceae): evidence for modulation by calcium pumps and apyrase activity.

PREMISE OF THE STUDY Gravity regulates the magnitude and direction of a trans-cell calcium current in germinating spores of Ceratopteris richardii. Blocking this current with nifedipine blocks the spores downward polarity alignment, a polarization that is fixed by gravity ∼10 h after light induces the spores to germinate. RNA-seq analysis at 10 h was used to identify genes potentially important for the gravity response. The data set will be valuable for other developmental and phylogenetic studies. METHODS De novo Newbler assembly of 958 527 reads from Roche 454 sequencing was executed. The sequences were identified and analyzed using in silico methods. The roles of endomembrane Ca(2+)-ATPase pumps and apyrases in the gravity response were further tested using pharmacological agents. KEY RESULTS Transcripts related to calcium signaling and ethylene biosynthesis were identified as notable constituents of the transcriptome. Inhibiting the activity of endomembrane Ca(2+)-ATPase pumps with 2,5-di-(t-butyl)-1,4-hydroquinone diminished the trans-cell current, but increased the orientation of the polar axis to gravity. The effects of applied nucleotides and purinoceptor antagonists gave novel evidence implicating extracellular nucleotides as regulators of the gravity response in these fern spores. CONCLUSIONS In addition to revealing general features of the transcriptome of germinating spores, the results highlight a number of calcium-responsive and light-receptive transcripts. Pharmacologic assays indicate endomembrane Ca(2+)-ATPases and extracellular nucleotides may play regulatory roles in the gravity response of Ceratopteris spores.

Electrical characterization of a single cell electroporation biochip with the 2-D scanning vibrating electrode technology

Advancements in microfabrication technology have lead to the development of planar micro-pore electroporation technology. This technology has been shown to provide greater control in single cell manipulation, and electroporation which is independent from cell size. In this work we report direct and spatially resolved characterization of electric currents within a planar micropore electroporation biochip to better understand this phenomenon at the cellular level. This work was performed using a two-dimensional (2-D) vibrating probe (VP). Analysis of the spatial patterns of current density yielded a 4th order polynomial profile in the planes parallel to the biochip’s surface and a three parameter hyperbolic decay profile in the planes perpendicular to the chip surface. A finite element model was developed which correlates with actual measurements on the micropore. Preliminary VP current density measurements of electroporated HepG2 cells revealed a significantly high current density minutes after electroporation even with non-electroporative pulses. These results indicate that cells take a considerable amount of time for complete electrophysiological recovery and indicate the use of the VP as a cell viability indicator for optimized electroporation.

Biochips and other microtechnologies for physiomics

This paper presents a review of microtechnologies relevant to applications in cellular physiology, including biochips, electrochemical sensors and optrodic sensing techniques. Microelectrodes have been the main tools for measuring cellular electrophysiology, oxygen, nitric oxide, neurotransmitters, pH and various ions. Optical fiber sensing methods, such as indicator-based optrodes, with fluorescence lifetime measurement, are now emerging as viable alternatives to electroanalytical chemistry. These new optrode techniques are possible because of recent advances in the optoelectronics industry and are comparably easier to miniaturize, have faster response times, do not consume the analyte and have lower operational costs. This review serves as a summary and predicts future trends for both electrochemical and optical luminescence lifetime sensing as components in lab-on-a-chip devices for physiological sensing.

Lab-on-a-Chip Approaches for Space-Biology Research

Lab-on-a-chip (LOC) systems with electrochemical sensing capability can provide real-time physiological measurements in spaceflight environments. They are easily miniaturized and integrated with existing space hardware systems. To reduce crew time during spaceflight research, the systems can be made autonomous and simple to use. Research and development of electrochemical-sensing LOC systems are still in progress for fundamental space-biology research in microgravity. Ion-selective electrodes as electrochemical sensors are miniaturized in an all-solid-state format for easier packaging and handling. The design, fabrication, and application of these sensors are discussed, with examples from those developed at the Physiological Sensing Facility (PSF) at Purdue University. The objective of this paper is not to provide an exhaustive review of current LOC systems, but to describe research developments made for the purpose of conducting physiological measurements in microgravity with examples of patents that support space missions.

Multi-analyte biochip (MAB) based on all-solid-state ion-selective electrodes (ASSISE) for physiological research.

Lab-on-a-chip (LOC) applications in environmental, biomedical, agricultural, biological, and spaceflight research require an ion-selective electrode (ISE) that can withstand prolonged storage in complex biological media (1-4). An all-solid-state ion-selective-electrode (ASSISE) is especially attractive for the aforementioned applications. The electrode should have the following favorable characteristics: easy construction, low maintenance, and (potential for) miniaturization, allowing for batch processing. A microfabricated ASSISE intended for quantifying H(+), Ca(2+), and CO3(2-) ions was constructed. It consists of a noble-metal electrode layer (i.e. Pt), a transduction layer, and an ion-selective membrane (ISM) layer. The transduction layer functions to transduce the concentration-dependent chemical potential of the ion-selective membrane into a measurable electrical signal. The lifetime of an ASSISE is found to depend on maintaining the potential at the conductive layer/membrane interface (5-7). To extend the ASSISE working lifetime and thereby maintain stable potentials at the interfacial layers, we utilized the conductive polymer (CP) poly(3,4-ethylenedioxythiophene) (PEDOT) (7-9) in place of silver/silver chloride (Ag/AgCl) as the transducer layer. We constructed the ASSISE in a lab-on-a-chip format, which we called the multi-analyte biochip (MAB) (Figure 1). Calibrations in test solutions demonstrated that the MAB can monitor pH (operational range pH 4-9), CO3(2-) (measured range 0.01 mM - 1 mM), and Ca(2+) (log-linear range 0.01 mM to 1 mM). The MAB for pH provides a near-Nernstian slope response after almost one month storage in algal medium. The carbonate biochips show a potentiometric profile similar to that of a conventional ion-selective electrode. Physiological measurements were employed to monitor biological activity of the model system, the microalga Chlorella vulgaris. The MAB conveys an advantage in size, versatility, and multiplexed analyte sensing capability, making it applicable to many confined monitoring situations, on Earth or in space. Biochip Design and Experimental Methods The biochip is 10 x 11 mm in dimension and has 9 ASSISEs designated as working electrodes (WEs) and 5 Ag/AgCl reference electrodes (REs). Each working electrode (WE) is 240 μm in diameter and is equally spaced at 1.4 mm from the REs, which are 480 μm in diameter. These electrodes are connected to electrical contact pads with a dimension of 0.5 mm x 0.5 mm. The schematic is shown in Figure 2. Cyclic voltammetry (CV) and galvanostatic deposition methods are used to electropolymerize the PEDOT films using a Bioanalytical Systems Inc. (BASI) C3 cell stand (Figure 3). The counter-ion for the PEDOT film is tailored to suit the analyte ion of interest. A PEDOT with poly(styrenesulfonate) counter ion (PEDOT/PSS) is utilized for H(+) and CO3(2-), while one with sulphate (added to the solution as CaSO4) is utilized for Ca(2+). The electrochemical properties of the PEDOT-coated WE is analyzed using CVs in redox-active solution (i.e. 2 mM potassium ferricyanide (K3Fe(CN)6)). Based on the CV profile, Randles-Sevcik analysis was used to determine the effective surface area (10). Spin-coating at 1,500 rpm is used to cast ~2 μm thick ion-selective membranes (ISMs) on the MAB working electrodes (WEs). The MAB is contained in a microfluidic flow-cell chamber filled with a 150 μl volume of algal medium; the contact pads are electrically connected to the BASI system (Figure 4). The photosynthetic activity of Chlorella vulgaris is monitored in ambient light and dark conditions.