Label-free plasmonic-based biosensing using a gold nanohole array chip coated with a wafer-scale deposited WS2 monolayer.

This paper reports the fabrication, testing and obtained performance of a plasmonic sensor employing a gold (Au) nanohole array chip coated with tungsten disulphide (WS2), which is then functionalized for the detection of protein-protein interactions. A key novelty is that the WS2 was deposited as a monoatomic layer using a wafer-scale synthesis method that successfully provided a film of both high quality and uniform thickness. The deposited WS2 film was transferred onto a Au nanohole array chip using a novel method and was subsequently functionalized with biotin. The final sensor was tested and it demonstrated efficient real-time and label-free plasmonic detection of biotin-streptavidin coupling. Specifically, compared to a standard (i.e. uncoated) Au nanohole-based sensor, our WS2-coated Au nanohole array boosted the spectral shift of the resonance wavelength by ∼190%, resulting in a 7.64-fold improvement of the limit of detection (LOD).

Low-Cost Method and Biochip for Measuring the Trans-Epithelial Electrical Resistance (TEER) of Esophageal Epithelium.

Trans-epithelial electrical resistance (TEER) is a good indicator of the barrier integrity of epithelial tissues and is often employed in biomedical research as an effective tool to assess ion transport and permeability of tight junctions. The Ussing chamber is the gold standard for measuring TEER of tissue specimens, but it has major drawbacks: it is a macroscopic method that requires a careful and labor intensive sample mounting protocol, allows a very limited viability for the mounted sample, has large parasitic components and low throughput as it cannot perform multiple simultaneous measurements, and this sophisticated and delicate apparatus has a relatively high cost. This paper demonstrates a low-cost home-made "sandwich ring" method which was used to measure the TEER of tissue specimens effectively. This method inspired the subsequent design of a biochip fabricated using standard soft lithography and laser engraving technologies, with which the TEER of pig epithelial tissues was measured. Moreover, it was possible to temporarily preserve the tissue specimens for days in the biochip and monitor the TEER continuously. Tissue responses after exposure tests to media of various pH values were also successfully recorded using the biochip. All these demonstrate that this biochip could be an effective, cheaper, and easier to use Ussing chamber substitute that may have relevant applications in clinical practice.

Recent advances in microfluidic methods in cancer liquid biopsy.

Early cancer detection, its monitoring, and therapeutical prediction are highly valuable, though extremely challenging targets in oncology. Significant progress has been made recently, resulting in a group of devices and techniques that are now capable of successfully detecting, interpreting, and monitoring cancer biomarkers in body fluids. Precise information about malignancies can be obtained from liquid biopsies by isolating and analyzing circulating tumor cells (CTCs) or nucleic acids, tumor-derived vesicles or proteins, and metabolites. The current work provides a general overview of the latest on-chip technological developments for cancer liquid biopsy. Current challenges for their translation and their application in various clinical settings are discussed. Microfluidic solutions for each set of biomarkers are compared, and a global overview of the major trends and ongoing research challenges is given. A detailed analysis of the microfluidic isolation of CTCs with recent efforts that aimed at increasing purity and capture efficiency is provided as well. Although CTCs have been the focus of a vast microfluidic research effort as the key element for obtaining relevant information, important clinical insights can also be achieved from alternative biomarkers, such as classical protein biomarkers, exosomes, or circulating-free nucleic acids. Finally, while most work has been devoted to the analysis of blood-based biomarkers, we highlight the less explored potential of urine as an ideal source of molecular cancer biomarkers for point-of-care lab-on-chip devices.

Microfluidic and Micromachined/MEMS Devices for Separation, Discrimination and Detection of Airborne Particles for Pollution Monitoring.

Most of the microfluidics-related literature describes devices handling liquids, with only a small part dealing with gas-based applications, and a much smaller number of papers are devoted to the separation and/or detection of airborne inorganic particles. This review is dedicated to this rather less known field which has become increasingly important in the last years due to the growing attention devoted to pollution monitoring and air quality assessment. After a brief introduction summarizing the main particulate matter (PM) classes and the need for their study, the paper reviews miniaturized devices and/or systems for separation, detection and quantitative assessment of PM concentration in air with portable and easy-to-use platforms. The PM separation methods are described first, followed by the key detection methods, namely optical (scattering) and electrical. The most important miniaturized reported realizations are analyzed, with special attention given to microfluidic and micromachined or micro-electro-mechanical systems (MEMS) chip-based implementations due to their inherent capability of being integrated in lab-on-chip (LOC) type of smart microsystems with increased functionalities that can be portable and are easy to use. The operating principles and (when available) key performance parameters of such devices are presented and compared, also highlighting their advantages and disadvantages. Finally, the most relevant conclusions are discussed in the last section.

Highlighting the uniqueness in dielectrophoretic enrichment of circulating tumor cells.

Circulating tumor cells (CTCs) play an essential role in the metastasis of tumors, and thus can serve as a valuable prognostic factor for malignant diseases. As a result, the ability to isolate and characterize CTCs is essential. This review underlines the potential of dielectrophoresis for CTCs enrichment. It begins by summarizing the key performance parameters and challenges of CTCs isolation using microfluidics. The two main categories of CTCs enrichment-affinity-based and label-free methods-are analysed, emphasising the advantages and disadvantages of each as well as their clinical potential. While the main argument in favour of affinity-based methods is the strong specificity of CTCs isolation, the major advantage of the label-free technologies is in preserving the integrity of the cellular membrane, an essential requirement for downstream characterization. Moving forward, we try to answer the main question: "What makes dielectrophoresis a method of choice in CTCs isolation?" The uniqueness of dielectrophoretic CTCs enrichment resides in coupling the specificity of the isolation process with the conservation of the membrane surface. The specificity of the dielectrophoretic method stems from the differences in the dielectric properties between CTCs and other cells in the blood: the capacitances of the malignantly transformed cellular membranes of CTCs differ from those of other cells. Examples of dielectrophoretic devices are described and their performance evaluated. Critical requirements for using dielectrophoresis to isolate CTCs are highlighted. Finally, we consider that DEP has the potential of becoming a cytometric method for large-scale sorting and characterization of cells.

An integrated on-chip platform for negative enrichment of tumour cells.

The study of cancer cells in blood, popularly called circulating tumour cells (CTCs), has exceptional prospects for cancer risk assessment and analysis. Separation and enrichment of CTCs by size-based methods suffer from a well-known recovery/purity trade-off while methods targeting certain specific surface proteins can lead to risk of losing CTCs due to Epithelial to Mesenchymal Transition (EMT) and thus adversely affect the separation efficiency. A negative selection approach is thus preferred for tumour cell isolation as it does not depend on biomarker expression or defines their physical property as the separation criteria. In this work, we developed a microfluidic chip to isolate CTCs from whole blood samples without targeting any tumour specific antigen. This chip employs a two-stage cell separation: firstly, magnetophoresis depletes the white blood cells (WBCs) from a whole blood sample and is then followed by a micro-slit membrane that enables depleting the red blood cells (RBCs) and retaining only the tumour cells. By creating strong magnetic field gradients along with customized antibody complexes to target WBCs, we are able to remove >99.9% of WBCs from 1:1 diluted blood at a sample processing rate of 500µL/min. This approach achieves an average of >80% recovery of spiked tumour cells from 2mL of whole blood in a total assay processing time of 50min without multiple processing steps.

Microfluidic immunomagnetic cell separation from whole blood.

Immunomagnetic-based separation has become a viable technique for the separation of cells and biomolecules. Here we report on the design and analysis of a simple and efficient microfluidic device for high throughput and high efficiency capture of cells tagged with magnetic particles. This is made possible by using a microfluidic chip integrated with customized arrays of permanent magnets capable of creating large magnetic field gradients, which determine the effective capturing of the tagged cells. This method is based on manipulating the cells which are under the influence of a combination of magnetic and fluid dynamic forces in a fluid under laminar flow through a microfluidic chip. A finite element analysis (FEA) model is developed to analyze the cell separation process and predict its behavior, which is validated subsequently by the experimental results. The magnetic field gradients created by various arrangements of magnetic arrays have been simulated using FEA and the influence of these field gradients on cell separation has been studied with the design of our microfluidic chip. The proof-of-concept for the proposed technique is demonstrated by capturing white blood cells (WBCs) from whole human blood. CD45-conjugated magnetic particles were added into whole blood samples to label WBCs and the mixture was flown through our microfluidic device to separate the labeled cells. After the separation process, the remaining WBCs in the elute were counted to determine the capture efficiency, and it was found that more than 99.9% WBCs have been successfully separated from whole blood. The proposed design can be used for positive selection as well as for negative enrichment of rare cells.

Microfluidic platform for negative enrichment of circulating tumor cells.

Negative enrichment is the preferred approach for tumor cell isolation as it does not rely on biomarker expression. However, size-based negative enrichment methods suffer from well-known recovery/purity trade-off. Non-size based methods have a number of processing steps that lead to compounded cell loss due to extensive sample processing and handling which result in a low recovery efficiency. We present a method that performs negative enrichment in two steps from 2 ml of whole blood in a total assay processing time of 60 min. This negative enrichment method employs upstream immunomagnetic depletion to deplete CD45-positive WBCs followed by a microfabricated filter membrane to perform chemical-free RBC depletion and target cells isolation. Experiments of spiking two cell lines, MCF-7 and NCI-H1975, in the whole blood show an average of >90 % cell recovery over a range of spiked cell numbers. We also successfully recovered circulating tumor cells from 15 cancer patient samples.

Label-free virus identification and characterization using electrochemical impedance spectroscopy.

We demonstrate here the application of electrochemical impedance spectroscopy (EIS) in microfluidic devices for label-free virus identification by means of their specific "signature" and also investigate its feasibility for titer quantitation using two basic approaches. The first one is a method based on identifying so-called "resonance" frequencies manifesting in our microdevices and monitoring their variation as a function of the virus concentration, whereas the second one relies on measuring the relative impedance variation at these "resonance" frequencies. Best results have been obtained for the highest "resonance" frequency (∼80 MHz), which we attribute to be due to both the structure of the microdevice and the extremely small size of the viruses that make their effect significant only at such frequencies. This is a simpler method of determining virus concentration in diluted solutions of purified viruses than the well-established traditional plaque assay titer estimation method, and-since it is based on frequency measurement-could potentially be more accurate.

Towards an optimal and unbiased approach for tumor cell isolation.

Our current understanding of clinical significance or the lack thereof of circulating tumor cells (CTCs) is biased by the technology used to isolate these rare cells. Despite the presence of a vast number of academic and commercial technologies, the lack of a standardized and optimized platform has been widely noted. We present a negative enrichment approach, integrating WBC depletion and chemical-free RBC depletion in the same setup without the need for centrifugation, washing or multiple sample handling steps. This approach achieves an average of >90 % recovery of spiked tumor cells and >99 % total WBC depletion in whole blood across multiple cell lines, in a simple and easy-to-use assay. The results presented herein and ongoing improvements aim to fulfill the need for a highly reliable, unbiased, standardized, and optimized CTC isolation platform, using component technologies that are validated for cell isolation.

Design and fabrication of Poly(dimethylsiloxane) arrayed waveguide grating.

We have designed, fabricated and characterized poly(dimethylsiloxane) (PDMS) arrayed waveguide grating (AWG) with four-channel output for operation in the visible light wavelength range. The PDMS AWG was realized based on the single-mode PDMS rib waveguide. The device was designed for 1 nm channel spacing with the wavelength ranging from 639 to 644 nm. The measured insertion loss is 11.4 dB at the peak transmission spectrum and the adjacent crosstalk is less than -16 dB. The AWG device occupies an area of 7.5 × 15 mm(2). PDMS AWG has the potential for integration with microfluidics in a monolithic PDMS lab-on-a-chip device for visible light spectroscopy applications.

Design and fabrication of poly(dimethylsiloxane) single-mode rib waveguide.

We have designed, fabricated and characterized poly(dimethylsiloxane) (PDMS) single-mode rib waveguides. PDMS was chosen specifically for the core and cladding. Combined with the soft lithography fabrication techniques, it enables an easy integration of microoptical components for lab-on-a-chip systems. The refractive index contrast, of 0.07% between the core and cladding for single-mode propagation was achieved by modifying the properties of the same base material. Alternatively, a higher refractive index contrast, of 1.18% was shown by using PDMS materials from two different manufacturers. The fabricated rib waveguides were characterized for mode profile characteristics and confirmed the excitation of the fundamental mode of the waveguide. The propagation loss of the single-mode rib waveguide was characterized using the cutback measurement method at a wavelength of 635 nm and found to be 0.48 dB/cm for of 0.07% and 0.20 dB/cm for of 1.18%. Y-branch splitter of PDMS single-mode rib waveguide was further demonstrated.

Algorithm and simulation for analysis of bio-images obtained by aperture diffraction based optical MEMS.

This paper proposes a novel method to detect transparent living cells in a transparent microfluidic chamber by optical diffraction of an aperture or an aperture array. Through the analysis of the far-field diffraction pattern, one of the parameters of the cells, including the size, refractive index, or position, can be extracted by the analysis software developed in this paper. Calculations are carried out to discuss the key issues of this MEMS device, and our simulation is verified by diffraction patterns of transparent microparticles on fabricated apertures, recorded via a digital camera.

Design of MEMS devices with optical apertures for the detection of transparent biological cells.

This paper provides a novel technique to detect transparent biological living cells trapped in a microfluidic MEMS device by optical diffraction. The device essentially consists of an optical aperture or an aperture array patterned in metal layer and a microfluidic chamber positioned above the center of the aperture. When the cells in the chamber are illuminated through the aperture, the far-field diffraction pattern can be recorded by a CCD camera or a photodetector array. This diffraction pattern uniquely corresponds to the sizes, positions, and intrinsic optical properties of the aperture, cells, and the microfluidic chamber materials, so any unknown but relevant parameter is able to be extrapolated when all other parameters are fixed or identified. This paper describes in detail the designs of various microfluidic chambers and apertures for this application, and the development of a complete set of software for the analysis of the cells' optical properties. Compared with other currently available methods for the detection of transparent living cells, this method has the advantages of simple device structure, easy to manipulate, able to simultaneously detect several cells of different species, as well as providing accurate and sensitive results. Besides the detection of living cells, this technique can also be used to detect or characterize other transparent or low optical absorption particles, such as polymer spheres or insoluble droplets, inside an aqueous solution.

Magnetic-based microfluidic platform for biomolecular separation.

A novel microfluidic platform for manipulation of micro/nano magnetic particles was designed, fabricated and tested for applications dealing with biomolecular separation. Recently, magnetic immunomagnetic cell separation has attracted a noticeable attention due to the high selectivity of such separation methods. Strong magnetic field gradients can be developed along the entire wire, and the miniaturized size of these current-carrying conductors strongly enhances the magnetic field gradient and therefore produces large, tunable and localized magnetic forces that can be applied on magnetic particles and confine them in very small spots. Further increases in the values of the generated magnetic field gradients can be achieved by employing miniaturized ferromagnetic structures (pillars) which can be magnetized by an external magnetic field or by micro-coils on the same chip. In this study, we demonstrate magnetic beads trapping, concentration, transportation and sensing in a liquid sample under continuous flow by employing high magnetic field gradients generated by novel multi-functional magnetic micro-devices. Each individual magnetic micro-device consists of the following components: 1. Cu micro-coils array embedded in the silicon substrate with high aspect ratio conductors for efficient magnetic field generation 2. Magnetic pillar(s) made of the magnetic alloy NiCoP for magnetic field focusing and magnetic field gradient enhancement. Each pillar is magnetized by its corresponding coil 3. Integrated sensing coil for magnetic beads detection 4. Microfluidic chamber containing all the previous components. Magnetic fields of about 0.1 T and field gradients of around 300 T/cm have been achieved, which allowed to develop a magnetic force of 3 x 10(-9) N on a magnetic particle with radius of 1 mum. This force is large enough to trap/move this particle as the required force to affect such particles in a liquid sample is on the order of approximately pN. Trapping rates of up to 80% were achieved. Furthermore, different micro-coil designs were realized which allowed various movement modes and with different step-sizes. These results demonstrate that such devices incorporated within a microfluidic system can provide significantly improved spatial resolution and force magnitude for quick, efficient and highly selective magnetic trapping, separation and transportation, and as such they are an excellent solution for miniaturized mu-total analysis systems.

An integrated microfluidic platform for magnetic microbeads separation and confinement.

An innovative microfluidic platform for magnetic beads manipulation is introduced, consisting of novel microfabricated 3D magnetic devices positioned in a microfluidic chamber. Each magnetic device comprises of an embedded actuation micro-coil in various design versions, a ferromagnetic pillar, a magnetic backside plate and a sensing micro-coil. The various designs of the micro-coils enable efficient magnetic beads trapping and concentration in different patterns. The finite element analysis (FEA) results show a significant increase of the developed force on suspended magnetic beads when the magnetic pillar and backside plate were integrated into the device structure. These simulation results were confirmed experimentally by measuring the magnetic beads trapping ratios for the different designs and structures of the devices under continuous flow conditions. The trapping ratios and profiles were studied using beads counting, measuring the change of inductance with the sensing micro-coil and by image processing. The devices have efficiently demonstrated a controlled and localized magnetic beads trapping and concentration at small spatial locations for the first time. The new results shown in this study demonstrate the feasibility of efficiently using these original devices as key elements in complex bio-analysis systems.