The ability to obtain sequence-specific genetic information about rare target organisms directly from complex biological samples at the point of care would transform many areas of biotechnology. Microfluidics technology offers compelling tools for integrating multiple biochemical processes in a single device, but despite significant progress, only limited examples have shown specific, genetic analysis of clinical samples within the context of a fully integrated, portable platform. Herein we present the Magnetic Integrated Microfluidic Electrochemical Detector (MIMED) that integrates sample preparation and electrochemical sensors in a monolithic disposable device to detect RNA-based virus directly from patient samples. By combining immunomagnetic target capture, concentration and purification, reverse-transcriptase polymerase chain reaction (RT-PCR) and single-stranded DNA (ssDNA) generation in the sample preparation chamber, as well as sequence specific electrochemical DNA detection in the electrochemical cell, we demonstrate the detection of influenza H1N1 in throat swab samples at loads as low as 10 TCID50 - 4 orders of magnitude below the clinical titer for this virus. Given the availability of affinity reagents for a broad range of pathogens, our system offers a general approach for multi-target diagnostics at the point-of-care.
Biological channels embedded in cell membranes regulate ionic transport by responding to external stimuli such as pH, voltage, and molecular binding. Mimicking the gating properties of these biological structures would be instrumental in the preparation of smart membranes used in biosensing, drug delivery, and ionic circuit construction. Here we present a new concept for building synthetic nanopores that can simultaneously respond to pH and transmembrane potential changes. DNA oligomers containing protonatable A and C bases are attached at the narrow opening of an asymmetric nanopore. Lowering the pH to 5.5 causes the positively charged DNA molecules to bind to other strands with negative backbones, thereby creating an electrostatic mesh that closes the pore to unprecedentedly high resistances of several tens of gigaohms. At neutral pH values, voltage switching causes the isolated DNA strands to undergo nanomechanical movement, as seen by a reversible current modulation. We provide evidence that the pH-dependent reversible closing mechanism is robust and applicable for nanopores with opening diameters of up to 14 nm. The concept of creating an electrostatic mesh may also be applied to different organic polymers.
A nanopore decorated with crown ether and DNA is selective to potassium ions over sodium ions at concentrations up to 1 M.
Effective systems for rapid, sequence-specific nucleic acid detection at the point of care would be valuable for a wide variety of applications, including clinical diagnostics, food safety, forensics, and environmental monitoring. Electrochemical detection offers many advantages as a basis for such platforms, including portability and ready integration with electronics. Toward this end, we report the Integrated Microfluidic Electrochemical DNA (IMED) sensor, which combines three key biochemical functionalities--symmetric PCR, enzymatic single-stranded DNA generation, and sequence-specific electrochemical detection--in a disposable, monolithic chip. Using this platform, we demonstrate detection of genomic DNA from Salmonella enterica serovar Typhimurium LT2 with a limit of detection of <10 aM, which is approximately 2 orders of magnitude lower than that from previously reported electrochemical chip-based methods.
Understanding the structure of aqueous electrolytes at interfaces is essential for predicting and optimizing device performance for a wide variety of emerging energy and environmental technologies. In this work, we investigate the structure of two common salt solutions, NaCl and KCl, at a hydrophobic interface within narrow carbon nanotubes (CNTs). Using a combination of first-principles and classical molecular dynamics simulations in conjunction with molecular orbital analysis, we find that the solvation structure of the cations in the CNTs can deviate substantially from the conventional weakly interacting hydrophobic picture. Instead, interactions between solvated ions and π orbitals of the CNTs are found to play a critically important role. Specifically, the ion solvation structure is ultimately determined by a complex interplay between cation−π interactions and the intrinsic flexibility of the solvation shell. In the case of K + , these effects result in an unusually strong propensity to partially desolvate and reside closer to the carbon wall than both Na + and Cl − , in sharp contrast with the known ion ordering at the water−vapor interface.
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