Biosensing in a microelectrofluidic system using optical whispering-gallery mode spectroscopy.
ABSTRACT: Label-free detection of biomolecules using an optical whispering-gallery mode sensor in a microelectrofluidic channel is simulated. Negatively charged bovine serum albumin is considered as the model protein analyte. The analyte transport in aqueous solution is controlled by an externally applied electrical field. The finite element method is employed for solving the equations of the charged species transport, the Poisson equation of electric potential, the equations of conservation of momentum and energy, and the Helmholtz equations of electromagnetic waves. The adsorption process of the protein molecules on the microsensor head surface is monitored by the resonance frequency shifts. Frequency shift caused by temperature variation due to Joule heating is analyzed and found to be negligible. The induced shifts behave in a manner similar to Langmuir-like adsorption kinetics; but the time constant increases due to the presence of the external electrical field. A correlation of the frequency shift, the analyte feed concentration in the solution, and the applied voltage gradient is obtained, in which an excellent linear relationship between the frequency shift and the analyte concentration is revealed. The applied voltage gradient enhances significantly the analyte concentration in the vicinity of the sensor surface; thus, the sensor sensitivity which has a power function of the voltage gradient with exponent 2.85 in the controlled voltage range. Simulated detection of extremely low protein concentration to the pico-molar level is carried out.
Project description:Colorimetry detects a color change resulted from a chemical reaction or molecular binding. Despite its widespread use in sensing, continuous monitoring of analytes with colorimetry is difficult, especially when the color-producing reaction or binding is irreversible. Here, we report on a gradient-based colorimetric sensor (GCS) to overcome this limitation. Lateral transport of analytes across a colorimetric sensor surface creates a color gradient that shifts along the transport direction over time, and GCS tracks the gradient shift and converts it into analyte concentration in real time. Using a low cost complementary metal-oxide semiconductor imager and imaging processing algorithm, we show submicrometer gradient shift tracking precision and continuous monitoring of ppb-level ozone.
Project description:We experimentally demonstrate dielectrophoretic concentration of biological analytes on the surface of a gold nanohole array, which concurrently acts as a nanoplasmonic sensor and gradient force generator. The combination of nanohole-enhanced dielectrophoresis, electroosmosis, and extraordinary optical transmission through the periodic gold nanohole array enables real-time label-free detection of analyte molecules in a 5 μL droplet using concentrations as low as 1 pM within a few minutes, which is more than 1000 times faster than purely diffusion-based binding. The nanohole-based optofluidic platform demonstrated here is straightforward to construct, applicable to both charged and neutral molecules, and performs a novel function that cannot be accomplished using conventional surface plasmon resonance sensors.
Project description:We apply polyelectrolyte multilayer films by consecutive alternate adsorption of positively charged polyallylamine hydrochloride and negatively charged sodium polystyrene sulfonate to the surface of graphene field effect transistors. Oscillations in the Dirac voltage shift with alternating positive and negative layers clearly demonstrate the electrostatic gating effect in this simple model system. A simple electrostatic model accounts well for the sign and magnitude of the Dirac voltage shift. Using this system, we are able to create p-type or n-type graphene at will. This model serves as the basis for understanding the mechanism of charged polymer sensing using graphene devices, a potentially technologically important application of graphene in areas such as DNA sequencing, biomarker assays for cancer detection, and other protein sensing applications.
Project description:Voltage sensing by voltage-gated sodium channels determines the electrical excitability of cells, but the molecular mechanism is unknown. beta-Scorpion toxins bind specifically to neurotoxin receptor site 4 and induce a negative shift in the voltage dependence of activation through a voltage sensor-trapping mechanism. Kinetic analysis showed that beta-scorpion toxin binds to the resting state, and subsequently the bound toxin traps the voltage sensor in the activated state in a voltage-dependent but concentration-independent manner. The rate of voltage sensor trapping can be fit by a two-step model, in which the first step is voltage-dependent and correlates with the outward gating movement of the IIS4 segment, whereas the second step is voltage-independent and results in shifted voltage dependence of activation of the channel. Mutations of Glu(779) in extracellular loop IIS1-S2 and both Glu(837) and Leu(840) in extracellular loop IIS3-S4 reduce the binding affinity of beta-scorpion toxin. Mutations of positively charged and hydrophobic amino acid residues in the IIS4 segment do not affect beta-scorpion toxin binding but alter voltage dependence of activation and enhance beta-scorpion toxin action. Structural modeling with the Rosetta algorithm yielded a three-dimensional model of the toxin-receptor complex with the IIS4 voltage sensor at the extracellular surface. Our results provide mechanistic and structural insight into the voltage sensor-trapping mode of scorpion toxin action, define the position of the voltage sensor in the resting state of the sodium channel, and favor voltage-sensing models in which the S4 segment spans the membrane in both resting and activated states.
Project description:The higher operating temperature of metal oxide and air instability of organic based NO2 sensor causes extremely urgent for development of a reliable low cost sensor to detect NO2 at room temperature. Therefore, we present a fabrication of large area Polymer/GO nano hybrid thin film for polymer thin film transistors (PTFTs) based NO2 sensors assisted via facile method named 'spreading-solidifying (SS) method', grown over air/liquid interface and successive investigation of effect of NO2 on film via several characterizations. The PTFTs sensor has demonstrated swift and high response towards low concentration of NO2 gas with air stability and provided real time non-invasive type NO2 sensor. Herein, we are reporting the nanohybrid PBTTT/GO composite based PTFT sensor with good repeatability and sensor response for low concentration NO2. The thin film grown via SS technique has reported very good adsorption/desorption of target analyte having response/recovery time of 75?s/523?s for 10 ppm concentration of NO2 gas. It has been observed that % change in drain current (sensor response) saturated with increasing concentration of NO2. The transient analysis demonstrates the fast sensor response and recovery time. Furthermore, in order to understand the insight of high performance of sensor, effect of NO2 on nanohybrid film and sensing mechanism, an in situ investigations was conducted via multiple technique viz. spectral, electronic, structural, and morphological characterization. Finally, the performance of sensor and the site of adsorption of NO2 at polymer chains were argued using schematic diagram. This work shows the simple fabrication process for mass production, low cost and room temperature operated gas sensors for monitoring the real-time environment conditions and gives an insight about the sensing mechanism adsorption site of NO2.
Project description:Synchronous neurotransmitter release is mediated by the opening of voltage-gated Ca(2+) channels and the build-up of submembrane Ca(2+) microdomains. Previous models of Ca(2+) microdomains have neglected possible electrostatic interactions between Ca(2+) ions and negative surface charges on the inner leaflet of the plasma membrane. To address the effects of these interactions, we built a computational model of ion electrodiffusion described by the Nernst-Planck and Poisson equations. We found that inclusion of a negative surface charge significantly alters the spatial characteristics of Ca(2+) microdomains. Specifically, close to the membrane, Ca(2+) ions accumulate, as expected from the strong electrostatic attraction exerted on positively charged Ca(2+) ions. Farther away from the membrane, increasing the surface charge density results in a reduction of the Ca(2+) concentration because of the preferential spread of Ca(2+) ions along lateral directions. The model also predicts that the negative surface charge will decrease the spatial gradient of the Ca(2+) microdomain in the lateral direction, resulting in increased overlap of microdomains originating from different Ca(2+) channels. Finally, we found that surface charge increases the probability of vesicle release if the Ca(2+) sensor is located within the electrical double layer, whereas this probability is decreased if the Ca(2+) sensor lies at greater distances from the membrane. Our data suggest that membrane surface charges exert a significant influence on the profile of Ca(2+) microdomains, and should be taken into account in models of neurotransmitter release.
Project description:Large conductance calcium- and voltage-sensitive K+ (MaxiK) channels share properties of voltage- and ligand-gated ion channels. In voltage-gated channels, membrane depolarization promotes the displacement of charged residues contained in the voltage sensor (S4 region) inducing gating currents and pore opening. In MaxiK channels, both voltage and micromolar internal Ca2+ favor pore opening. We demonstrate the presence of voltage sensor rearrangements with voltage (gating currents) whose movement and associated pore opening is triggered by voltage and facilitated by micromolar internal Ca2+ concentration. In contrast to other voltage-gated channels, in MaxiK channels there is charge movement at potentials where the pore is open and the total charge per channel is 4-5 elementary charges.
Project description:A dual-analyte fluorescent chemosensor (ExoSensor 517) for the direct visualization of neurotransmitters released upon exocytosis is presented. The sensor exploits the high concentration of neurotransmitters (e.g., glutamate, norepinephrine, and dopamine) and the pH gradient between the vesicle and synaptic cleft. The cooperative recognition elements require both binding and a change in environmental pH to afford a fluorescence response which makes ExoSensor 517 one of the first integrated molecular logic gates to be used for biological applications.
Project description:The position and extent of movement of a charged peptide within a membrane bilayer provides much controversy. In our study, we have examined the nature of the highly charged helix-turn-helix motif (S3b and S4) to address how a highly charged peptide is stabilized within a bilayer in the presence of various transmembrane electrical potentials. Our double-bilayer simulation results show how the variation of the salt concentrations between the inner and outer bath establishes a transmembrane potential. Our results also show that important features of the peptide affected by changes in electrical potential are the center of mass depth, the swivel/kink degrees of conformation, and the hydrogen-bonding patterns. As the voltage gradient across the bilayer increased, the center of mass of the peptide shifted in a direction toward the outer bath. The peptide also has a higher percent helical content and the swivel/kink conformation is more rigid for nonpolarized systems where no voltage drop occurred between salt baths. Our results also provide some suggestions for how this domain may be affected by environmental changes as part of the voltage sensor in a K-channel.
Project description:Niflumic acid, 2-[[3-(trifluoromethyl)phenyl]amino]pyridine-3-carboxylic acid (NFA), is a nonsteroidal anti-inflammatory drug that also blocks or modifies the gating of many ion channels. Here, we investigated the effects of NFA on hyperpolarization-activated cyclic nucleotide-gated cation (HCN) pacemaker channels expressed in X. laevis oocytes using site-directed mutagenesis and the two-electrode voltage-clamp technique. Extracellular NFA acted rapidly and caused a slowing of activation and deactivation and a hyperpolarizing shift in the voltage dependence of HCN2 channel activation (-24.5 +/- 1.2 mV at 1 mM). Slowed channel gating and reduction of current magnitude was marked in oocytes treated with NFA, while clamped at 0 mV but minimal in oocytes clamped at -100 mV, indicating the drug preferentially interacts with channels in the closed state. NFA at 0.1 to 3 mM shifted the half-point for channel activation in a concentration-dependent manner, with an EC(50) of 0.54 +/- 0.068 mM and a predicted maximum shift of -38 mV. NFA at 1 mM also reduced maximum HCN2 conductance by approximately 20%, presumably by direct block of the pore. The rapid onset and state-dependence of NFA-induced changes in channel gating suggests an interaction with the extracellular region of the S4 transmembrane helix, the primary voltage-sensing domain of HCN2. Neutralization (by mutation to Gln) of any three of the outer four basic charged residues in S4, but not single mutations, abrogated the NFA-induced shift in channel activation. We conclude that NFA alters HCN2 gating by interacting with the extracellular end of the S4 voltage sensor domains.