Smartphone-interfaced lab-on-a-chip devices for field-deployable enzyme-linked immunosorbent assay

The emerging technologies on mobile-based diagnosis and bioanalytical detection have enabled powerful laboratory assays such as enzyme-linked immunosorbent assay (ELISA) to be conducted in field-use lab-on-a-chip devices. In this paper, we present a low-cost universal serial bus (USB)-interfaced mobile platform to perform microfluidic ELISA operations in detecting the presence and concentrations of BDE-47 (2,2′,4,4′-tetrabromodiphenyl ether), an environmental contaminant found in our food supply with adverse health impact. Our point-of-care diagnostic device utilizes flexible interdigitated carbon black electrodes to convert electric current into a microfluidic pump via gas bubble expansion during electrolytic reaction. The micropump receives power from a mobile phone and transports BDE-47 analytes through the microfluidic device conducting competitive ELISA. Using variable domain of heavy chain antibodies (commonly referred to as single domain antibodies or Nanobodies), the proposed device is sensitive for a BDE-47 concentration range of 10⁻³–10⁴μg/l, with a comparable performance to that uses a standard competitive ELISA protocol. It is anticipated that the potential impact in mobile detection of health and environmental contaminants will prove beneficial to our community and low-resource environments.     

Supernatant decanting on a centrifugal platform

This study presents a novel approach to decant supernatant on a centrifugal platform. By manipulating the centrifugally induced pressure and the elastic deformation of the plastic lids in the decanting chamber, fixed amounts of the supernatant can be decanted into the detection chamber at lower rotational speeds. The experimental results showed that decanted volume is affected by the volume of deformation and the operating parameters. Factors that influence the decanting ratio are also discussed. This approach has the advantages of simple design and low manufacturing cost; further, it has no need of surface modification. It has been applied to on-disk separation of plasma from whole blood, and the results showed good stability and repeatability.     

A negative-pressure-driven microfluidic chip for the rapid detection of a bladder cancer biomarker in urine using bead-based enzyme-linked immunosorbent assay

This paper describes an integrated microfluidic chip that is capable of rapidly and quantitatively measuring the concentration of a bladder cancer biomarker, apolipoprotein A1, in urine samples. All of the microfluidic components, including the fluid transport system, the micro-valve, and the micro-mixer, were driven by negative pressure, which simplifies the use of the chip and facilitates commercialization. Magnetic beads were used as a solid support for the primary antibody, which captured apolipoprotein A1 in patients'' urine. Because of the three-dimensional structure of the magnetic beads, the concentration range of the target that could be detected was as high as 2000 ng ml⁻¹. Because this concentration is 100 times higher than that quantifiable using a 96-well plate with the same enzyme-linked immunosorbent assay (ELISA) kit, the dilution of the patient''s urine can be avoided or greatly reduced. The limit of detection was determined to be approximately 10 ng ml⁻¹, which is lower than the cutoff value for diagnosing bladder cancer (11.16 ng ml⁻¹). When the values measured using the microfluidic chip were compared with those measured using conventional ELISA using a 96-well plate for five patients, the deviations were 0.9%, 6.8%, 9.4%, 1.8%, and 5.8%. The entire measurement time is 6-fold faster than that of conventional ELISA. This microfluidic device shows significant potential for point-of-care applications.     

Adsorption and isolation of nucleic acids on cellulose magnetic beads using a three-dimensional printed microfluidic chip

While advances in genomics have enabled sensitive and highly parallel detection of nucleic acid targets, the isolation and extraction of the nucleic acids remain a critical bottleneck in the workflow. We present here a simple 3D printed microfluidic chip that allows for the vortex and centrifugation free extraction of nucleic acids. This novel microfluidic chip utilizes the presence of a water and oil interface to filter out the lysate contaminants. The pure nucleic acids, while bound on cellulose particles, are magnetically moved across the oil layer. We demonstrated efficient and rapid extraction of spiked Human Papillomavirus (HPV) 18 plasmids in specimen transport medium, in under 15 min. An overall extraction efficiency of 61% is observed across a range of HPV plasmid concentrations (5 × 10¹ to 5 × 10⁶ copies/100 μl). The magnetic, interfacial, and viscous drag forces inside the microgeometries of the chip are modeled. We have also developed a kinetics model for the adsorption of nucleic acids on cellulose functionalized superparamagnetic beads. We also clarify here the role of carrier nucleic acids in the adsorption and isolation of nucleic acids. Based on the various mechanistic insights detailed here, customized microfluidic devices can be designed to meet the range of current and emerging point of care diagnostics needs.     

Resistive pulse sensing of magnetic beads and supraparticle structures using tunable pores

Tunable pores (TPs) have been used for resistive pulse sensing of 1 μm superparamagnetic beads, both dispersed and within a magnetic field. Upon application of this field, magnetic supraparticle structures (SPSs) were observed. Onset of aggregation was most effectively indicated by an increase in the mean event magnitude, with data collected using an automated thresholding method. Simulations enabled discrimination between resistive pulses caused by dimers and individual particles. Distinct but time-correlated peaks were often observed, suggesting that SPSs became separated in pressure-driven flow focused at the pore constriction. The distinct properties of magnetophoretic and pressure-driven transport mechanisms can explain variations in the event rate when particles move through an asymmetric pore in either direction, with or without a magnetic field applied. Use of TPs for resistive pulse sensing holds potential for efficient, versatile analysis and measurement of nano- and microparticles, while magnetic beads and particle aggregation play important roles in many prospective biosensing applications.     

Experimental validation of numerical study on thermoelectric-based heating in an integrated centrifugal microfluidic platform for polymerase chain reaction amplification

A comprehensive study involving numerical analysis and experimental validation of temperature transients within a microchamber was performed for thermocycling operation in an integrated centrifugal microfluidic platform for polymerase chain reaction (PCR) amplification. Controlled heating and cooling of biological samples are essential processes in many sample preparation and detection steps for micro-total analysis systems. Specifically, the PCR process relies on highly controllable and uniform heating of nucleic acid samples for successful and efficient amplification. In these miniaturized systems, the heating process is often performed more rapidly, making the temperature control more difficult, and adding complexity to the integrated hardware system. To gain further insight into the complex temperature profiles within the PCR microchamber, numerical simulations using computational fluid dynamics and computational heat transfer were performed. The designed integrated centrifugal microfluidics platform utilizes thermoelectrics for ice-valving and thermocycling for PCR amplification. Embedded micro-thermocouples were used to record the static and dynamic thermal responses in the experiments. The data collected was subsequently used for computational validation of the numerical predictions for the system response during thermocycling, and these simulations were found to be in agreement with the experimental data to within ∼97%. When thermal contact resistance values were incorporated in the simulations, the numerical predictions were found to be in agreement with the experimental data to within ∼99.9%. This in-depth numerical modeling and experimental validation of a complex single-sided heating platform provide insights into hardware and system design for multi-layered polymer microfluidic systems. In addition, the biological capability along with the practical feasibility of the integrated system is demonstrated by successfully performing PCR amplification of a Group B Streptococcus gene.     

Specific binding and magnetic concentration of CD8+ T-lymphocytes on electrowetting-on-dielectric platform

In the quest to create a low-power portable lab-on-a-chip system, we demonstrate the specific binding and concentration of human CD8+ T-lymphocytes on an electrowetting-on-dielectric (EWOD)-based digital microfluidic platform using antibody-conjugated magnetic beads (MB-Abs). By using a small quantity of nonionic surfactant, we enable the human cell-based assays with selective magnetic binding on the EWOD device in an air environment. High binding efficiency (～92%)of specific cells on MB-Abs is achieved due to the intimate contact between the cells and the magnetic beads (MBs) produced by the circulating flow within the small droplet. MBs have been used and cells manipulated in the droplets actuated by EWOD before; reported here is a cell assay of a clinical protocol on the EWOD device in air environment. The present technique can be further extended to capture other types of cells by suitable surface modification on the MBs.     

A multiplexed immunoaggregation biomarker assay using a two-stage micro resistive pulse sensor

We present an immunoaggregation assay chip for multiplexed biomarkers detection. This chip is based on immunoaggregation of antibody functionalized microparticles (Ab-MPs) to quantify concentrations of multiple biomarkers simultaneously. A mixture of multiple types of Ab-MPs probes with different sizes and magnetic properties, which were functionalized by different antibodies, was used for the multiplexed assay. The interactions between biomarkers and their specific Ab-MPs probes caused the immunoaggregation of Ab-MPs. A two-stage micro resistive pulse sensor was used to differentiate and count the Ab-MP aggregates triggered by different biomarkers via size and magnetic property for multiplexed detection. The volume fraction of each type of Ab-MP aggregates indicates the concentration of the corresponding target biomarker. In our study, we demonstrated multiplexed detection of two model biomarkers (human ferritin and mouse anti-rabbit IgG) in 10% fetal bovine serum, using anti-ferritin Ab and anti-mouse IgG Ab functionalized MPs. We found that the volume fraction of Ab-MP aggregates increased with the increased biomarker concentrations. The detection ranges from 5.2 ng/ml to 208 ng/ml and 3.1 ng/ml to 5.12 × 10⁴ng/ml were achieved for human ferritin and mouse anti-rabbit IgG. This bioassay chip is able to quantitatively detect multiple biomarkers in a single test without fluorescence or enzymatic labeling process and hence is promising to serve as a useful tool for rapid detection of multiple biomarkers in biomedical research and clinical applications.     

Estimation of the levels of atrial natriuretic peptide (ANP) in cardiac blood samples by a simple and indigenous enzyme linked immunosorbent assay

Levels of Atrial Natriuretic Peptide (ANP) were estimated in twenty four blood samples collected from the right and the left ventricles of the patients undergoing cardiac catheterization by an Enzyme Linked Immunosorbent Assay (ELISA) developed to the sensitivity of 5 pp/well and within 10% of interassay coeficient of variance. Simultaneously, levels of ANP in plasma samples from the systemic venous blood of seventy five normal subjects were also estimated which ranged between 25–60 pg/ml.     

Serum arylsulfatase A assay in metachromatic leukodystrophy: An experience in a neuropsychiatric set-up

Metachromatic leukodystrophy is a lysosomal disease caused mainly by a deficiency of the enzyme arylsulfatase A. The assay of arylsulfatase A in the serum provides a fast and easy method for the confirmatory diagnosis of this disorder. Serum arylsulfatase A was estimated in 52 normal healthy control subjects and 269 patients with symptoms of cerebral white matter disease in order to diagnose and confirm metachromatic leukodystrophy. A total of eight cases of metachromatic leukodystrophy with a low serum arylsulfatase A was detected, of which three cases were of the late-infantile type, four cases the juvenile type and only one case the adult type.     

The role of pre-innovation platform activity for diffusion success: Evidence from consumer innovations on a 3D printing platform

Digital platforms are becoming increasingly important for household sector innovators that seek support for the innovation process and that want to make innovations available to large audiences. Innovation development and diffusion is especially challenging for first-time innovators as they cannot build on experiences from prior innovations. We argue that first-time innovators can increase the diffusion success of their innovations by engaging in pre-innovation platform activities. We use the context of the 3D printing platform Thingiverse to show that a consumer's pre-innovation platform activity increases innovation diffusion success and that frequency, quality and relatedness of a consumer's pre-innovation platform activity promotes this effect. We find support that innovation quality, the use of recombinant innovations, and innovation documentation are three mechanisms through which pre-innovation platform activities translate into higher diffusion success of consumers’ first innovation.     

Dual-mode on-demand droplet routing in multiple microchannels using a magnetic fluid as carrier phase

We present dual-mode, on-demand droplet routing in a multiple-outlet microfluidic device using an oil-based magnetic fluid. Magnetite (Fe₃O₄) nanoparticle-contained oleic acid (MNOA) was used as a carrier phase for droplet generation and manipulation. The water-in-MNOA droplets were selectively distributed in a curved microchannel with three branches by utilizing both a hydrodynamic laminar flow pattern and an external magnetic field. Without the applied magnetic field, the droplets travelled along a hydrodynamic centerline that was displaced at each bifurcating junction. However, in the presence of a permanent magnet, they were repelled from the centerline and diverted into the desired channel when the repelled distance exceeded the minimum offset allocated to the channel. The repelled distance, which is proportional to the magnetic field gradient, was manipulated by controlling the magnet''s distance from the device. To evaluate routing performance, three different sizes of droplets with diameters of 63, 88, and 102 μm were directed into designated outlets with the magnet positioned at varying distances. The result demonstrated that the 102-μm droplets were sorted with an accuracy of ∼93%. Our technique enables on-demand droplet routing in multiple outlet channels by simply manipulating magnet positions (active mode) as well as size-based droplet separation with a fixed magnet position (passive mode).     

Isolation of cells for selective treatment and analysis using a magnetic microfluidic chip

This study describes the development and testing of a magnetic microfluidic chip (MMC) for trapping and isolating cells tagged with superparamagnetic beads (SPBs) in a microfluidic environment for selective treatment and analysis. The trapping and isolation are done in two separate steps; first, the trapping of the tagged cells in a main channel is achieved by soft ferromagnetic disks and second, the transportation of the cells into side chambers for isolation is executed by tapered conductive paths made of Gold (Au). Numerical simulations were performed to analyze the magnetic flux and force distributions of the disks and conducting paths, for trapping and transporting SPBs. The MMC was fabricated using standard microfabrication processes. Experiments were performed with E. coli (K12 strand) tagged with 2.8 μm SPBs. The results showed that E. coli can be separated from a sample solution by trapping them at the disk sites, and then isolated into chambers by transporting them along the tapered conducting paths. Once the E. coli was trapped inside the side chambers, two selective treatments were performed. In one chamber, a solution with minimal nutrition content was added and, in another chamber, a solution with essential nutrition was added. The results showed that the growth of bacteria cultured in the second chamber containing nutrient was significantly higher, demonstrating that the E. coli was not affected by the magnetically driven transportation and the feasibility of performing different treatments on selectively isolated cells on a single microfluidic platform.     

基于JQuery Mobile的国际漫游自助服务平台设计与实现

随着移动互联网技术的快速发展,JQuery Mobile框架以其成本低廉、兼容性强、跨平台、使用简单等特性越来越受到移动开发人员的欢迎。文章结合移动用户投诉国际漫游中遇到的实际问题,基于JQuery Mobile技术设计开发了国际漫游自助服务平台。通过使用该平台,有效降低了用户国际漫游相关问题处理时长,提升了用户满意度。     

An extract-then-abstract based method to generate disaster-news headlines using a DNN extractor followed by a transformer abstractor

Generating news headlines has been one of the predominant problems in Natural Language Processing research. Modern transformer models, if fine-tuned, can present a good headline with attention to all the parts of a disaster-news article. A disaster-news headline generally focuses on the event, its effect, and the major impacts, which a transformer model lacks when generating the headline. The extract-then-abstract based method proposed in this article improves the performance of a state-of-the-art transformer abstractor to generate a good-quality disaster-news headline. In this work, a Deep Neural Network (DNN) based sentence extractor and a transformer-based abstractive summarizer work sequentially to generate a headline. The sentence extraction task is formulated as a binary classification problem where the DNN model is trained to generate two binary labels: one corresponding to the sentence similarity with ground truth headlines and the other corresponding to the presence of disaster and its impact related information in the sentence. The transformer model generates the headline from the sentences extracted by the DNN. ROUGE scores of the headlines generated using the proposed method are found to be better than the scores of the headlines generated directly from the original documents. The highest ROUGE 1, 2, and 3 score improvements are observed in the case of the Text-To-Text Transfer Transformer (T5) model by 17.85%, 38.13%, and 21.01%, respectively. Such improvements suggest that the proposed method can have a high utility for finding effective headlines from disaster related news articles.     

Rare cell isolation and profiling on a hybrid magnetic/size-sorting chip

We present a hybrid magnetic/size-sorting (HMSS) chip for isolation and molecular analyses of circulating tumor cells (CTCs). The chip employs both negative and positive cell selection in order to provide high throughput, unbiased CTC enrichment. Specifically, the system utilizes a self-assembled magnet to generate high magnetic forces and a weir-style structure for cell sorting. The resulting device thus can perform multiple functions, including magnetic depletion, size-selective cell capture, and on-chip molecular staining. With such capacities, the HMSS device allowed one-step CTC isolation and single cell detection from whole blood, tested with spiked cancer cells. The system further facilitated the study of individual CTCs for heterogeneity in molecular marker expression.Circulating tumor cells (CTCs) have emerged as an important biomarker in clinical practice as well as in fundamental research.^{1, 2} CTCs, shed from primary tumors, have been shown to be an early harbinger of tumor expansion and metastasis³ and have been used to predict disease progression, response to treatment, relapse, and overall survival.^{4, 5, 6} Recent work has shown that CTCs display distinct proteomic and genetic profiles; for example, CTCs in pancreatic cancer, have increased RNA expression of Wnt, implicating this pathway in metastasis.⁷ Proteomic characterization of proliferative markers such as Ki-67, and hormonal markers such as androgen receptor in prostate cancer, also have been shown to be predictive of treatment outcome.^{8, 9}Despite such clinical potential of CTCs, their routine detection and characterization still remains a significant technical challenge.¹⁰ The task requires screening of a large number of cells (e.g., > 10⁷ cells in 10 ml blood) and enrichment of heterogeneous targets against a complex biological background. Two main methods of CTC isolation are typically used: positive and negative selection. In positive selection, CTCs are directly isolated from blood via size-based filtration^{11, 12, 13, 14, 15, 16, 17, 18, 19, 20} or antibody-based capture.^{1, 8, 21} Negative depletion reduces abundant blood cells, often by immunomagnetic separation, for downstream CTC enrichment.²² Both approaches have been used for high throughput CTC isolation from whole blood (SI Table 1).²³ Each method, however, has its own inherent limitations. Positive enrichment could be biased by its selection criteria (e.g., cell size and cell surface markers). Negative selection, albeit unbiased, often requires complex sample processing (e.g., multiple washing steps for CTC isolation) that could result in cell loss.We hypothesized that both positive and negative selection could be combined in a single platform to enable (1) highly efficient and unbiased CTC purification, and (2) in-situ molecular analyses of collected cells. As a proof-of-concept, we herein describe a hybrid magnetic/size-sorting (HMSS) system that integrates magnetic and size-based isolation into a compact microfluidic chip. The HMSS first uses a magnetic filter to deplete leukocytes through immunomagnetic capture. Samples then pass through a size-sorter region that traps individual cells at predefined locations. Since abundant leukocytes are removed by the magnetic filter, the size-sorter could have a low size cut-off (∼5 μm), which allows for the unbiased capture of even small cancer cells. Furthermore, molecular probes can be introduced to perform on-chip, multiplexed analyses at single-cell resolution. We evaluated the utility of the developed system by capturing and profiling tumor cells in whole blood. The HMSS offers the advantages of both negative and positive selection and thereby differs from the recently reported iChip system²⁴ which can operate only in either a negative or a positive selection mode.