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.

Effects of the electrode size and modification protocol on a label-free electrochemical biosensor.

In the present work, the effect of a surface modification protocol along with the electrode size has been investigated for developing an efficient, label-free electrochemical biosensing method for diagnosis of traumatic brain injury (TBI) biomarkers. A microdisk electrode array (MDEA) and a macroelectrode with a comb structure (MECS) were modified with an anti-GFAP (GFAP = glial fibrillary acidic protein) antibody using two protocols for optimum and label-free detection of GFAP, a promising acute-phase TBI biomarker. For the MDEA, an array of six microdisks with a 100 µm diameter and, for the MECS, a 3.2 mm × 5.5 mm electrode 5 µm wide with 10 µm spaced comb fingers were modified using an optimized protocol for dithiobis(succinimidyl propionate) (DSP) self-assembled monolayer formation. Anti-GFAP was covalently bound, and the remaining free DSP groups were blocked using ethanolamine (Ea). Sensors were exposed to solutions with different GFAP concentrations, and a label-free electrochemical impedance spectroscopy (EIS) technique was used to determine the concentration. EIS results confirmed that both types of Ea/anti-GFAP/DSP/Au electrodes modified with an optimized DSP-based protocol can accurately detect GFAP in the range of 1 pg mL(-1) to 100 ng mL(-1) with a detection limit of 1 pg mL(-1). However, the cross-use of the MDEA protocol on the MECS and vice versa resulted in very low sensitivity or poor signal resolution, underscoring the importance of proper matching of the electrode size and type and the surface modification protocol.

Electrically controlled giant piezoresistance in silicon nanowires.

Herein we demonstrate giant piezoresistance in silicon nanowires (NWs) by the modulation of an electric field-induced with an external electrical bias. Positive bias for a p-type device (negative for an n-type) partially depleted the NWs forming a pinch-off region, which resembled a funnel through which the electrical current squeezed. This region determined the total current flowing through the NWs. In this report, we combined the electrical biasing with the application of mechanical stress, which impacts the charge carriers' concentration, to achieve an electrically controlled giant piezoresistance in nanowires. This phenomenon was used to create a stress-gated field-effect transistor, exhibiting a maximum gauge factor of 5000, 2 orders of magnitude increase over bulk value. Giant piezoresistance can be tailored to create highly sensitive mechanical sensors operating in a discrete mode such as nanoelectromechanical switches.

The electromechanical response of silicon nanowires to buckling mode transitions.

Here we show how the electromechanical properties of silicon nanowires (NWs) are modified when they are subjected to extreme mechanical deformations (buckling and buckling mode transitions), such as those appearing in flexible devices. Flexible devices are prone to frequent dynamic stress variations, especially buckling, while the small size of NWs could give them an advantage as ultra-sensitive electromechanical stress sensors embedded in such devices. We evaluated the NWs post-buckling behavior and the effects of buckling mode transition on their piezoresistive gauge factor (GF). Polycrystalline silicon NWs were embedded in SiO(2) microbridges to facilitate concurrent monitoring of their electrical resistance without problematic interference, while an external stylus performed controlled deformations of the microbridges. At points of instability, the abrupt change in the buckling configuration of the microbridge corresponded to a sharp resistance change in the embedded NWs, without altering the NWs' GF. These results also highlight the importance of strategically positioning the NW in the devices, since electrical monitoring of buckling mode transitions is feasible when the deformations impact a region where the NW is placed. The highly flexible NWs also exhibited unusually large fracture strength, sustaining tensile strains up to 5.6%; this will prove valuable in demanding flexible sensors.

Engineered biochar via microwave CO2 and steam pyrolysis to treat carcinogenic Congo red dye.

We developed an innovative single-step pyrolysis approach that combines microwave heating and activation by CO2 or steam to transform orange peel waste (OPW) into microwave activated biochar (MAB). This involves carbonization and activation simultaneously under an inert environment. Using CO2 demonstrates dual functions in this approach, acting as purging gas to provide an inert environment for pyrolysis while activating highly porous MAB. This approach demonstrates rapid heating rate (15-120 °C/min), higher temperature (> 800 °C) and shorter process time (15 min) compared to conventional method using furnace (> 1 h). The MAB shows higher mass yield (31-44 wt %), high content of fixed carbon (58.6-61.2 wt %), Brunauer Emmett Teller (BET) surface area (158.5-305.1 m2/g), low ratio of H/C (0.3) and O/C (0.2). Activation with CO2 produces more micropores than using steam that generates more mesopores. Steam-activated MAB records a higher adsorption efficiency (136 mg/g) compared to CO2 activation (91 mg/g), achieving 89-93 % removal of Congo Red dye. The microwave pyrolysis coupled with steam or CO2 activation thereby represents a promising approach to transform fruit-peel waste to microwave-activated biochar that remove hazardous dye.

Solid phase nucleic acid extraction technique in a microfluidic chip using a novel non-chaotropic agent: dimethyl adipimidate.

Here, we present a silicon microfluidic system for the purification and extraction of nucleic acids from human body fluid samples utilizing a dimethyl adipimidate (DMA)-based solid-phase extraction method. We propose DMA, which has been used as an amino-reactive cross-linking agent within cells and proteins, as a non-chaotropic reagent for the capture of nucleic acids to overcome the limitations of existing chaotropic and non-chaotropic techniques such as low binding efficiency, PCR inhibition and so on. DMA contains bi-functional imidoesters that form reversible cross-linking structures with DNA therefore providing a high surface-area to volume ratio for capturing DNA without structurally modifying microfluidic channels. In this work, we have first demonstrated highly efficient capture and purification of genomic DNA (T24 cell line) with DMA using a label-free silicon microring resonator sensor device. In addition, we observed the improvement of the DNA amplification efficiency by using the proposed technique for both the genetic (HRAS) and epigenetic (RARß) analysis of DNA biomarkers. Particularly, we confirmed that the DMA-based solid-phase extraction technique can be applied for the extraction of genomic DNA with higher purity (p < 0.001) using human body fluids (blood and urine) in silicon microfluidic devices compared to other chaotropic methods. Therefore, the proposed technique would be able to harmonize with a micro-total analysis system platform for the analysis of genetic and epigenetic DNA biomarkers related to human diseases in the field of point-of-care (POC) diagnostic applications.

Lab-on-a-chip technology: impacting non-invasive prenatal diagnostics (NIPD) through miniaturisation.

This paper aims to provide a concise review of non-invasive prenatal diagnostics (NIPD) to the lab-on-a-chip and microfluidics community. Having a market of over one billion dollars to explore and a plethora of applications, NIPD requires greater attention from microfluidics researchers. In this review, a complete overview of conventional diagnostic procedures including invasive as well as non-invasive (fetal cells and cell-free fetal DNA) types are discussed. Special focus is given to reviewing the recent and past microfluidic approaches to NIPD, as well as various commercial entities in NIPD. This review concludes with future challenges and ethical considerations of the field.

EIS-based biosensor for ultra-sensitive detection of TNF-α from non-diluted human serum.

Serum background is a critical issue for biosensor development as it interferes with the detection of target molecules and may give rise to false positive signal. We present here highly sensitive and selective TNF-α biosensor which is able to detect TNF-α from non-diluted human serum using magnetic bead coupled antibody and electrochemical impedance spectroscopy (EIS) techniques. The process is designed to detect TNF-α from human serum in three stages; (1) abundant protein backgrounds are depleted from the serum using magnetic bead coupled albumin and IgG antibodies, (2) after background depletion TNF-α is captured using magnetic bead coupled TNF-α antibody, and (3) the captured TNF-α is eluted from the magnetic beads and measured using EIS technique in which comb structured gold microelectrodes array (CSGM) is utilized to enhance the detection sensitivity. The system is able to achieve the limit of detection (LOD) at 1 pg/ml (57 fM) and a linear relationship between increasing TNF-α concentrations and charge-transfer resistance in a dynamic range of 1-1000 pg/ml.

Anti-EpCAM modified LC-SPDP monolayer on gold microelectrode based electrochemical biosensor for MCF-7 cells detection.

A succinimidyl 6-(3-[2-pyridyldithio]-propionamido) hexanoate (LC-SPDP) self-assembled monolayer (SAM) prepared onto a 500 µm (diameter) gold microelectrode (Au) surface has been utilized for covalent immobilization of anti-EpCAM antibody. Amino group on anti-EpCAM antibody was covalently bound with succinimidyl group on SAM via amide bond and unreacted active groups of LC-SPDP were blocked using 1% ethanol amine (EA). These anti-EpCAM/LC-SPDP/Au electrodes were characterized using cyclic voltammetric (CV) and fluorescence techniques, respectively. The anti-EpCAM/LC-SPDP/Au electrodes were exposed to solutions with different MCF-7 cell concentrations and CV technique was used to determine the cell concentration. Further, CV studies on blank 500 and 50 µm (diameter) gold microelectrodes were used to identify cell via molecular profiling using ferrocene amidopentyl carboxylic acid based redox tagging and magnetic beads based enhancement. CV results confirm that the anti-EpCAM/LC-SPDP/Au based biosensor could detect MCF-7 cells in the range of 1×10(5)-1×10(8) with correlation coefficient of 0.999 and detection limit of 1×10(5) cells ml(-1) i.e. 100 cells in solution used for incubation (1 µl). Molecular profiling studies suggest that smaller size microelectrode (50 µm; diameter) with magnetic beads based enhancement can be employed to identify cell type. This work establishes the feasibility of using microelectrode based platform for breast cancer specific MCF-7 cell concentration estimation and their molecular profiling.

Size based sorting and patterning of microbeads by evaporation driven flow in a 3D micro-traps array.

We present a three-dimensional (3D) micro-traps array for size selective sorting and patterning of microbeads via evaporation-driven capillary flow. The interconnected micro-traps array was manufactured by silicon micromachining. Microliters of aqueous solution containing particle mixtures of different sized (0.2 to 20 µm diameter) beads were dispensed onto the micro-traps substrate. The smaller particles spontaneously wicked towards the periphery of the chip, while the larger beads were orderly docked within the micro-traps array.

Fabrication of self-sealed circular nano/microfluidic channels in glass substrates.

We realized self-sealing fluidics channels with circular cross-sections having diameters ranging between 30 and 2000 nm on a 200 mm glass wafer through CMOS compatible processes. Lateral voids were narrowed and sealed with non-conformal plasma enhanced chemical vapour deposition (PECVD) of phospho silicate glass (PSG) along silicon oxide trenches on silicon wafers. Leveraging on the reflow properties of PSG, circular profiled-channels were formed after undergoing high temperature annealing. These devices were subsequently transferred onto a borosilicate glass substrate through anodic bonding, and a fully transparent microfluidic device was achieved with the complete removal of the handle silicon substrate. The process offers a means of integrating electrochemical and optical sensing on the same platform, for biological research.