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Inspired by electronic transistors, electric field gating has been adopted to manipulate ionic currents of smart nanofluidic devices. Here, we report a PNP nanofluidic bipolar junction transistor (nBJT) consisting of one polyaniline (PANI) layer sandwiched between two polyethylene terephthalate (PET) nanoporous membranes. The PNP nBJT exhibits three different responses of currents (quasi-linear, rectification, and sigmoid) due to the counterbalance between surface charge distribution and base voltage applied in the nanofluidic channels; thus, they can be switched by base voltage. Four operating modes (cutoff, active, saturation, and breakdown mode) occur in the collector response currents. Under optimal conditions, the PNP nBJT exhibits an average current gain of up to 95 in 100 mM KCl solution at a low base voltage of 0.2 V. The present nBJT is promising for fabrication of nanofluidic devices with logical-control functions for analysis of single molecules.

Biological ion pumps with two separate gates can actively transport ions against the concentration gradient. Developing an artificial nanofluidic device with multiple responsive sites is of great importance to improve its controllability over ion transport to further explore its logic function and mimic the biological process. Here, we propose an electrochemical polymerization method to fabricate electrochemically switchable double-gate nanofluidic devices. The ion transport of the double-gate nanofluidic device can be in situ and reversibly switched among four different states. The logic function of this nanofluidic device is systematically investigated by assuming the gate state as the input and the transmembrane ionic conductance as the output. A biomimetic electrochemical ion pump is then established by alternately applying two different specific logic combinations, realizing an active ion transport under a concentration gradient. This work would inspire further studies to construct complex logical networks and explore bioinspired ion pump systems.

Micro-/nanofluidics has received considerable attention over the past two decades, which allows efficient biomolecule trapping and preconcentration due to ion concentration polarization (ICP) within nanostructures. The rich scientific content related to ICP has been widely exploited in different applications including protein concentration, biomolecules sensing and detection, cell analysis, and water purification. Compared to pure microfluidic devices, micro-/nanofluidic devices show a highly efficient sample enrichment capacity and nonlinear electrokinetic flow feature. These two unique characterizations make the micro-/nanofluidic systems promising in high-performance bioanalysis. This review provides a comprehensive description of the ICP phenomenon and its applications in bioanalysis. Perspectives are also provided for future developments and directions of this research field.

We propose an in situ and label-free method for detection of biomolecular recognition events by use of a nanochannel-ion channel hybrid device integrated with an electrochemical detector. The aptamer is first immobilized on the outer surface of the nanochannel-ion channel hybrid. Its binding with target thrombin in solution considerably regulates the mass-transfer behavior of the device owing to the varied surface charge density and effective channel size. Via the electrochemical detector, the changed mass-transport property can be monitored in real time, which enables in situ and label-free detection of thrombin-aptamer recognition. The solution pH has a significant influence on detection sensitivity. Under optimal pH conditions, a detection limit as low as 0.22 fM thrombin can be achieved, which is much lower than most reported work. The present nanofluidic device provides a simple, ultrasensitive, and label-free platform for monitoring biomolecular recognition events, which would hold great potential in exploring the functions and reaction mechanisms of biomolecules in living systems.

Single cell analysis is essential for understanding the heterogeneity, behaviors of cells, and diversity of target analyte in different subcellular regions. Nucleolin (NCL) is a multifunctional protein that is markedly overexpressed in most of the cancer cells. The variant expression levels of NCL in subcellular regions have a marked influence on cancer proliferation and treatments. However, the specificity of available methods to identify the cancer biomarkers is limited because of the high level of subcellular matrix effect. Herein, we proposed a novel technique to increase both the molecular and spectral specificity of cancer diagnosis by using aptamers affinity based portable nanopipette with distinctive surface-enhanced Raman scattering (SERS) activities. The aptamers-functionalized gold-coated nanopipette was used to capture target, while p-mercaptobenzonitrile (MBN) and complementary DNA modified Ag nanoparticles (AgNPs) worked as Raman reporter to produce SERS signal. The SERS signal of Raman nanotag was lost upon NCL capturing via modified DNA aptamers on nanoprobe, which further helped to verify the specificity of nanoprobe. For proof of concept, NCL protein was specifically extracted from different cell lines by aptamers modified SERS active nanoprobe. The nanoprobes manifested specifically good affinity for NCL with a dissociation constant K d of 36 nM and provided a 1000-fold higher specificity against other competing proteins. Furthermore, the Raman reporter moiety has a vibrational frequency in the spectroscopically silent region (1800-2300 cm -1 ) with a negligible matrix effect from cell analysis. The subcellular localization and spatial distribution of NCL were successfully achieved in various types of cells, including MCF-7A, HeLa, and MCF-10A cells. This type of probing technique for single cell analysis could lead to the development of a new perspective in cancer diagnosis and treatment at the cellular level.

The sensitive identification of single nucleotide polymorphisms becomes increasingly important for disease diagnosis, prevention, and practical applicability of pharmacogenomics. Herein, we propose a simple, highly selective, label-free single nucleotide polymorphisms (SNPs) sensing device by electrochemically monitoring the diffusion flux of ferricyanide probe across probe DNA/morpholino duplex functionalized nanochannels of porous anodic alumina. When perfectly matched or mismatched target DNA flows through the nanochannels modified with probe DNA/morpholino duplex, it competes for the probe DNA from morpholino, resulting in a change of the surface charges. Thus, the diffusion flux of negatively charged electroactive probe ferricyanide is modulated since it is sensitive to the surface charge due to the electrostatic interactions in electric double layer-merged nanochannels. Monitoring of the change in diffusion flux of probe enables us to detect not only a single base or two base mismatched sequence but also the specific location of the mismatched base. As is demonstrated, SNPs in the PML/RARα fusion gene, known as a biomarker of acute promyelocytic leukemia (APL), have been successfully detected.

Exploring local pH in micro/nanoscale is fundamentally important for understanding microprocesses including the corrosion of metal and the metabolism of cell. Regular fluorescence pH probes and potentiometric electrodes show either low signal intensity or lack of spatial resolution when being applied in a micro/nanoenvironment. Here, we developed a nanoscale reversible pH probe based on the plasmonic coupling effect of i-motif modulated gold nanoparticle (AuNP) assembly. The pH probe shows a reversible and highly sensitive response to pH variation between 4.5 and 7.5. Introduction of morpholino oligomers (MO), a neutral analog of DNA, into the assembly endows the pH probe with high stability even under low salt concentration. The intense optical signal of a AuNP enables local pH to be read out not only in the micro/nanofluidic channel but also on a single i-motif-MO-AuNP assembly. Recording of the strong plasmonic resonance scattering spectrum of AuNP provides a promising method for extracting chemical information in nanospace of biological systems.

A nanochannel array based nanodevice can mimic the biological environments and thus unveil the natural properties, conformation and recognition information of biomolecules such as proteins and DNA in confined spaces. Here we report that porous anodic alumina (PAA) of a highly parallel nanochannel array covalently modified with proteins significantly modulates the transport of a negatively charged probe of ferricyanide due to the electrostatic interactions between the probes and modified nanochannel inner surface. Results show that such electrostatic interaction exists in a wide range of ionic strength from 1 mM to 100 mM in 20 nm nanochannels modified with proteins (hemoglobin, bovine serum albumin, and goat anti-rabbit IgG secondary antibody). In addition, the maximal steady-state flux of the charged probe through the modified nanochannel array is directly related to the ionic strength which determines the electric double layer thickness and solution pH which modulates the nanochannel surface charge. Thus, the modulated mass transport of the probe by solution pH can be used to study the charge properties of the immobilized proteins in nanochannel confined conditions, leading us to obtain the isoelectric point (pI) of the proteins confined in nanochannels. The determined pI values of two known proteins of hemoglobin and bovine serum albumin are close to the ones of the same proteins covalently modified on a 3-mercaptopropionic acid self-assembled monolayer/gold electrode. In addition, the pI of an unknown protein of goat anti-rabbit IgG secondary antibody confined in nanochannels was determined to be 6.3. Finally, the confinement effect of nanochannels on the charge properties of immobilized proteins has been discussed.

The nanochannel (in a porous layer) and ion-channel (in a barrier layer) hybrid structure of anodic alumina is used as a protein-trapping device. The transmembrane potential drives the electromigration of the charged proteins (FITC-labeled) into the nanochannels, but electromigration across the barrier layer is impossible due to the size-exclusion effect. As a result, the proteins can be continuously trapped in the nanochannels., (Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

This paper presents a simple and cost-effective UV-ablation technique for fabrication of size-tunable nanofluidics devices via photochemical decomposition reaction. UV-irradiation through a PET photomask results in continuous decomposition of poly(carbonate) (PC), forming nanochannel and carboxyl groups on the surface of the etched PC. This photochemical decomposition process occurs at molecular scale, therefore, the depth of nanochannels can be controlled at nanometer level. The etching rate is estimated to be ca. 0.015 nms(-1). To demonstrate the potential application of the present UV-ablation technique, a nanochannel was fabricated and integrated with microchannels to form a micro/nanofluidics chip for protein concentration. Using this device, about 10(3)-10(5) fold protein concentration can be achieved within 10 min. The present approach offers a simple and practical solution to fabricate nanofluidics devices at low-cost, and the resulting device could provide ideal platforms for μTAS towards various applications in biology and chemistry., (Copyright © 2011 Elsevier B.V. All rights reserved.)

A strategy for label-free oligonucleotide (DNA) analysis has been proposed by measuring the DNA-morpholino hybridization hindered diffusion flux of probe ions Fe(CN)(6)(3-) through nanochannels of a porous anodic alumina (PAA) membrane. The flux of Fe(CN)(6)(3-) passing through the PAA nanochannels is recorded using an Au film electrochemical detector sputtered at the end of the nanochannels. Hybridization of the end-tethered morpholino in the nanochannel with DNA forms a negatively charged DNA-morpholino complex, which hinders the diffusion of Fe(CN)(6)(3-) through the nanochannels and results in a decreased flux. This flux is strongly dependent on ionic strength, nanochannel aperture, and target DNA concentration, which indicates a synergetic effect of steric and electrostatic repulsion effects in the confined nanochannels. Further comparison of the probe flux with different charge passing through the nanochannels confirms that the electrostatic effect between the probe ions and DNA dominates the hindered diffusion process. Under optimal conditions, the present nanochannel array-based DNA biosensor gives a detection limit of 0.1 nM.

In this paper, a micro/nanofluidic preconcentration device integrated with an electrochemical detector has been used to study the enrichment of enzymes and homogeneous enzyme reaction kinetics. The enzymes are first concentrated in front of a nanochannel via an exclusion-enrichment effect (EEE) mechanism of the nanochannel integrated in a microfluidics device. If a substrate is electrokinetically transported to the concentrated enzymes, homogeneous enzymatic reaction occurs. The enzymatic reaction product can penetrate through the nanochannel to be detected electrochemically. In this device, the enriched enzymes can be well retained and repeatedly used, thus, the enzymatic reaction occurs in a continuous-flow mode. For demonstration, Glucose oxidase (GOx) was chosen as the model enzyme to study the influence of enzyme concentration on its reaction kinetics. The different concentration of GOx in front of the nanochannel was simply achieved by using different enrichment time. When substrate glucose was introduced electrokinetically, a rapid electrochemical steady-state response could be obtained. It was found that the electrochemical response to a constant glucose concentration increased with the increase of enzyme enrichment time, which is expected for homogeneous enzymatic reactions. Under proper conditions, the electrochemical responds linearly to the glucose concentration ranging from 0 to 15 mM, and the Michaelis constants (K(m)) are relatively low, which indicates a more efficient complex formation between enzyme and substrate. These results suggest that the present micro/nanofluidics device is promising for the study of enzymatic reaction kinetics and other bioassays such as cell assays, drug discovery, and clinical diagnosis.

1D PB nanowires with good electrochemical activities have been prepared by using DNA molecules as the templates. By using the photosynthesis method, PB nanowires were successfully synthesized in solution and on smooth surface, respectively. The PB nanowires are made of many connected spherical PB nanoparticles with the average diameter of about 10 nm at a relative short UV radiation time. Upon increasing the reaction time, the PB nanoparticles grow into zigzag nanowires, and finally form a network of long chain structures. The morphologies and properties of the PB nanomaterials were characterized by using TEM, IR, UV-vis measurements. In addition, the resultant PB nanowires have good electrochemical activities in comparing with the PB nanoparticles prepared without DNA templates. The present easy method could be interesting for the construction of electrochromic and biosensing devices.

We used polyethyleneglycol (PEG) as a modulator to manipulate pore and cell sizes in the porous anodic alumina (PAA) fabrication. It is shown for the first time that continuous manipulation of the pore size of PAA can be realized. Combined with the coexistent cell-size controlling effect, the morphology and properties of this important nanoscale template and separation membrane can be precisely regulated. The pore size modulation mechanism is proposed on the basis of the morphological and electrochemical results. The presence of PEG in the electrolyte results in a more compacted structure of the barrier layer alumina (BLA), although the barrier layer thickness does not change considerably. In addition, the additive can obviously restrain the chemical dissolution of alumina and shape smaller pores. These two effects combined with the increased viscosity of the electrolyte slow down the ion transportation and diminish the anodization current. Thus, the burning-down phenomena of the aluminum substrates can be avoided at relatively high voltage anodization, and an interpore distance up to 610 nm can be achieved.

We developed a facile and rapid one-step technique for design and fabrication of passive micromixers in microfluidic devices using a direct-printing process. A laser printing mechanism was dexterously adopted to pattern the microchannels with different gray levels using vector graphic software. With the present method, periodically ordered specific bas-relief microstructures can be easily fabricated on transparencies by a simple printing process. The size and shape of the resultant microstructures are determined by the gray level of the graphic software and the resolution of the laser printer. Patterns of specific bas-relief microstructures on the floor of a channel act as obstacles in the flow path for advection mixing, which can be used as efficient mixing elements. The mixing effect of the resultant micromixer in microfluidic devices was evaluated using CCD fluorescence spectroscopy. We found that the mixing performance depends strongly on the gray level values. Under optimal conditions, fast passive mixing with our periodic ordered patterns in microfluidic devices has been achieved at the very early stages of the laminar flow. In addition, fabrication of micromixers using the present versatile technique requires less than an hour. The present method is promising for fabrication of micromixers in microfluidic devices at low cost and without complicated devices and environment, providing a simple solution to mixing problems in the micro-total-analysis-systems field.