Microchip Technology Research Articles

Microfluidic Sample Preparation for Medical Diagnostics.

Fast and reliable diagnoses are invaluable in clinical care. Samples (e.g., blood, urine, and saliva) are collected and analyzed for various biomarkers to quickly and sensitively assess disease progression, monitor response to treatment, and determine a patient's prognosis. Processing conventional samples entails many manual time-consuming steps. Consequently, clinical specimens must be processed by skilled technicians before antigens or nucleic acids are detected, and these are often present at dilute concentrations. Recently, several automated microchip technologies have been developed that potentially offer many advantages over traditional bench-top extraction methods. The smaller length scales and more refined transport mechanisms that characterize these microfluidic devices enable faster and more efficient biomarker enrichment and extraction. Additionally, they can be designed to perform multiple tests or experimental steps on one integrated, automated platform. This review explores the current research on microfluidic methods of sample preparation that are designed to aid diagnosis, and covers a broad spectrum of extraction techniques and designs for various types of samples and analytes.

Printed Flexible Plastic Microchip for Viral Load Measurement through Quantitative Detection of Viruses in Plasma and Saliva.

We report a biosensing platform for viral load measurement through electrical sensing of viruses on a flexible plastic microchip with printed electrodes. Point-of-care (POC) viral load measurement is of paramount importance with significant impact on a broad range of applications, including infectious disease diagnostics and treatment monitoring specifically in resource-constrained settings. Here, we present a broadly applicable and inexpensive biosensing technology for accurate quantification of bioagents, including viruses in biological samples, such as plasma and artificial saliva, at clinically relevant concentrations. Our microchip fabrication is simple and mass-producible as we print microelectrodes on flexible plastic substrates using conductive inks. We evaluated the microchip technology by detecting and quantifying multiple Human Immunodeficiency Virus (HIV) subtypes (A, B, C, D, E, G, and panel), Epstein-Barr Virus (EBV), and Kaposi’s Sarcoma-associated Herpes Virus (KSHV) in a fingerprick volume (50 µL) of PBS, plasma, and artificial saliva samples for a broad range of virus concentrations between 102 copies/mL and 107 copies/mL. We have also evaluated the microchip platform with discarded, de-identified HIV-infected patient samples by comparing our microchip viral load measurement results with reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) as the gold standard method using Bland-Altman Analysis.

A microfluidic device for label-free, physical capture of circulating tumor cell clusters.

Cancer cells metastasize through the bloodstream either as single migratory circulating tumor cells (CTCs) or as multicellular groupings (CTC-clusters). Existing technologies for CTC enrichment are designed primarily to isolate single CTCs, and while CTC-clusters are detectable in some cases, their true prevalence and significance remain to be determined. Here, we developed a microchip technology (Cluster-Chip) specifically designed to capture CTC-clusters independent of tumor-specific markers from unprocessed blood. CTC-clusters are isolated through specialized bifurcating traps under low shear-stress conditions that preserve their integrity and even two-cell clusters are captured efficiently. Using the Cluster-Chip, we identify CTC-clusters in 30–40% of patients with metastatic cancers of the breast, prostate and melanoma. RNA sequencing of CTC-clusters confirms their tumor origin and identifies leukocytes within the clusters as tissue-derived macrophages. Together, the development of a device for efficient capture of CTC-clusters will enable detailed characterization of their biological properties and role in cancer metastasis.

A parallel implementation of an off-lattice individual-based model of multicellular populations

As computational models of multicellular populations include ever more detailed descriptions of biophysical and biochemical processes, the computational cost of simulating such models limits their ability to generate novel scientific hypotheses and testable predictions. While developments in microchip technology continue to increase the power of individual processors, parallel computing offers an immediate increase in available processing power. To make full use of parallel computing technology, it is necessary to develop specialised algorithms. To this end, we present a parallel algorithm for a class of off-lattice individual-based models of multicellular populations. The algorithm divides the spatial domain between computing processes and comprises communication routines that ensure the model is correctly simulated on multiple processors. The parallel algorithm is shown to accurately reproduce the results of a deterministic simulation performed using a pre-existing serial implementation. We test the scaling of computation time, memory use and load balancing as more processes are used to simulate a cell population of fixed size. We find approximate linear scaling of both speed-up and memory consumption on up to 32 processor cores. Dynamic load balancing is shown to provide speed-up for non-regular spatial distributions of cells in the case of a growing population.

The Design of EMG Measurement System for Arm Strength Training Machine

The setup of interactive communication between arm strength training machine and the people will make exercise and rehabilitation therapy become more friendly. The employment of electromyographic not only can help physical therapy but also can achieve more effective rehabilitation. Both of the system hardware and software of the arm strength training machine with EMG system are well designed and described. The fundamental design of electromyographic measurement system based on a microcontroller is analyzed and discussed. The software programming is developed in MPLAB integrated development environment from the Microchip Technology Inc. and the friendly user interface is created as well. Finally, an arm strength training machine with electromyographic control system is realized and demonstrated. The experimental results show the feasibility and fidelity of the complete designed system.

High Performance Nanostructured Silicon Oxide Impedance Biosensor System with Online Noise Spectroscopy Analysis

In this paper, noise spectroscopy has been reported earlier as an efficient tool for enhancing the selectivity of biosensors and the analysis was carried out by desktop spectrum analyzer. There were no reports, however, on the efficacy of this method in complex mixture. We have performed here for the first time, noise spectroscopy analysis on complex mixture of food toxin samples and observed that the first cut-off frequencies are indicative of the fact whether the solution has only specific antigen, mixture of specific and nonspecific antigen or no specific antigen at all. To realize a portable immunosensor, we have developed an electronic interface using digital signal processor (DSP) chip of Microchip Technology which has the embedded Fast Fourier Transform (FFT) algorithm for computation of noise spectrum. The entire system has been successfully demonstrated to detect 0.1 fg/ml aflatoxin B1 (AfB1) from a complex mixture with as high as 1000ng/ml non-specific toxins.

Ki-1/57 and CGI-55 ectopic expression impact cellular pathways involved in proliferation and stress response regulation

Ki-1/57 (HABP4) and CGI-55 (SERBP1) are regulatory proteins and paralogs with 40.7% amino acid sequence identity and 67.4% similarity. Functionally, they have been implicated in the regulation of gene expression on both the transcriptional and mRNA metabolism levels. A link with tumorigenesis is suggested, since both paralogs show altered expression levels in tumor cells and the Ki-1/57 gene is found in a region of chromosome 9q that represents a haplotype for familiar colon cancer. However, the target genes regulated by Ki-1/57 and CGI-55 are unknown. Here, we analyzed the alterations of the global transcriptome profile after Ki-1/57 or CGI-55 overexpression in HEK293T cells by DNA microchip technology. We were able to identify 363 or 190 down-regulated and 50 or 27 up-regulated genes for Ki-1/57 and CGI-55, respectively, of which 20 were shared between both proteins. Expression levels of selected genes were confirmed by qRT-PCR both after protein overexpression and siRNA knockdown. The majority of the genes with altered expression were associated to proliferation, apoptosis and cell cycle control processes, prompting us to further explore these contexts experimentally. We observed that overexpression of Ki-1/57 or CGI-55 results in reduced cell proliferation, mainly due to a G1 phase arrest, whereas siRNA knockdown of CGI-55 caused an increase in proliferation. In the case of Ki-1/57 overexpression, we found protection from apoptosis after treatment with the ER-stress inducer thapsigargin. Together, our data give important new insights that may help to explain these proteins putative involvement in tumorigenic events.

MOLECULAR CHARACTERIZATION OF MYCOBACTERIUM TUBERCULOSIS STRAINS CIRCULATING IN THE URAL REGION, RUSSIA

Abstract. Overall 178 Mycobacterium tuberculosis isolates recovered in 2009–2011 from newly diagnosed epidemiologically unlinked to TB patients in the Ural region of Russia have been studied. The absolute concentration method was used for drug susceptibility testing. Mutations katG, inhA and rpoB associated with resistance to isoniazid and rifampicin were detected by microchip technology («TB-Biochip»). The isolates were genotyped by real-time PCR for the detection of Beijing/non-Beijing genotypes and 15-locus MIRU-VNTR typing according to «MIRU-VNTRplus» (http://www.miru-vntrplus.org). More than half (55.1%) of 178 isolates belonged to the Beijing family, 58.7% of them were multiple drug resistant (MDR) mostly due to rpoBSer531→Leu and katGSer315→Thr1 substitutions. Fifty VNTR profiles were found in 98 Beijing isolates; 57 of them grouped into 9 clusters. The largest VNTR cluster included 23 (23.5%) Beijing isolates and 21 of them were MDR. The 80 non-Beijing isolates showed 64 distinct VNTR patterns which belonged to 6 genetic families: LAM, Ural, Haarlem, etc. Among LAM and Ural isolates 30.4% and 28.6% were MDR, respectively. The 5 of 7 MDR LAM isolates had specific mutation profile: rpoBAsp516→Val substitution and mutations katGSer315→Thr1 and inhA_T15. The MDR Ural isolates showed the heterogeneity of mutations in rpoB gene compared to other genotypes. Taken together, these findings suggest the emergence and spread of MDR-TB in the Ural region which is characterized by circulation of MDR strains of different genotypes with the Beijing family genotype to be predominant.

A Remote Monitoring System for Voltage, Current, Power and Temperature Measurements

Abstract This paper presents a study and design of a monitoring system for the continuous measurement of electrical energy parameters such as voltage, current, power and temperature. This system is designed to monitor the data remotely over internet. The electronic power meter is based on a microcontroller from Microchip Technology Inc. PIC family. The design takes into consideration the correct operation in the event of an outage or brown out by recording the electrical values and the temperatures in EEPROM internally available in the microcontroller. Also a digital display is used to show the acquired measurements. A computer will remotely monitor the data over internet.

Lab on a Microchip and Microfluidic Technologies: Toxology and Drug Development

LAB-ON-A-CHIP Ann Heslin Picture the length of two centimeters. Now, picture a synthetic chip that can mimic all functions of an entire human lung within this two-centimeter length. Pretty remarkable, right? What is even more remarkable is that this in vitro (existing outside a living organism) device is precise on a micro-scale (10-6) and has been shown to be a reliable human lung model. This lung-on-a-chip device is one example of a broader category of synthetic devices known as ‘lab- on-a chip’ or ‘microfluidic’ devices. Lab-on-a-chip is what it implies: miniaturized science. Though the theory of miniaturization is not hard to conceptualize, the process of turning theory into practice has proven to be much harder. Never- theless, meticulous engineering has led to the rapid advance of these devices and has paved an auspicious path for their contribution to organ modeling. One challenge the lung model faced was how to recreate a functional alveolar-capillary interface that is present in all human lungs. To understand this model, let us briefly revisit the physiology of a lung. During respiration, air passes down through the lungs into alveolar sacs where gas exchange occurs (Fig 1). But before oxygen can enter the bloodstream, it first diffuses through alveolar epithelial cells and then passes through a fused basement membrane, where it encounters the capillary endothelial cells. In the lung model, a 10 μm porous mem- brane composed of poly(dimethylsiloxane) (PDMS) represented the fused basement membrane, which had epithelial and endothelial cells growing as B S J Figure 1. monolayers on opposing side (Fig 2). Extracellular matrix (ECM), which can be thought of as a gluing substance, was coated on both sides of the PDMS membrane to allow the two cell lines to firmly adhere. Once attached to the material, both cell lines grew to 100 percent confluence (the whole PDMS membrane was covered with cells) and sustained themselves for a prolonged period of over two weeks. Next, this thin PDMS membrane was clamped between two thicker layers to form two smaller side chambers and one larger central cavity (Fig 3). An etching solution was pumped through the two side microchannels so that only the larger, central cavity had an upper and lower compartment. This engineering allowed for air to enter the upper compartment, mimicking the air-liquid inter- face within alveoli. A vascular medium (blood-like substance) was also allowed to run simultaneously through the lower compartment to emulate capillary blood-flow. The lung model also accounted for the physical movements—expansion and contraction—that occur during respiration. During inspiration, the diaphragm contracts (moves downwards) to decrease the intra- pleural pressure inside the alveolar sacs (sacs expand in volume), which increases the efficiency of gas ex- change (Fig 1). A vacuum was applied to the two side micro chambers to create a pressure-driven stretching of the thin PDMS membrane (Fig 4). This vacuum-in- duced stretching distorted the cultured epithelial and endothelial cells, forcing them to survive a physical environment similar to one that occurs in vivo (inside a living organism). Figure 2. 28 • B erkeley S cientific J ournal • S ynthetics • S pring 2014 • V olume 18 • I ssue 2

Microchip electrophoresis–copper nanowires for fast and reliable determination of monossacharides in honey samples

Microchip electrophoresis (ME) with electrochemical detection has been demonstrated to be a powerful tool in food analysis. However, the coupling of ME with electrochemical detection and nanotechnologies is still in its infancy, knowing that nanomaterials can significantly improve the ME analytical performance. This work reports the coupling between ME and copper nanowires (CuNWs) for the selective analysis of monosaccharides in honey samples. Also, in terms of real applicability, the study of analytical reliability of ME is an issue of paramount importance. To this end, a representative group of nine honey samples were analyzed and the results were compared with those previously obtained by HPLC-refractive index. ME-CuNWs approach allowed the separation of glucose and fructose in <250 s under optimized separation (20 mM NaOH + 10 mM H3 BO3 , pH 12; separation voltage + 1000 V) and detection (E = +0.70 V in 20 mM NaOH + 10 mM H3 BO3 , pH 12) conditions. An excellent stability of EOF during sample analysis was achieved with RSDs for migration times <2% and for amperometric currents <9%. The quantitative contents for individual glucose and fructose obtained using ME-CuNWs in comparison with those obtained by HPLC-refractive index were highly in agreement with errors <10% indicating the reliability of the approach. The excellent analytical performance obtained confirms the analytical potency of ME-CuNWs approach, enhancing the maturity of the microchip technology and opening new avenues for future implementation of applications in the field of food analysis.

Copper Nanowires Immobilized on the Boards of Microfluidic Chips for the Rapid and Simultaneous Diagnosis of Galactosemia Diseases in Newborn Urine Samples

Galactosemia is a rare disease that is diagnosed through the identification of different metabolite profiles. Therefore, the specific detection of galactose 1-phosphate (Gal 1-P), galactose (Gal), and uridyl diphosphate galactose (UDP-Gal) confirms type I, II, and III galactosemia diseases. Because of the low prevalence of galactosemia, sample availability is very scarce and screening methods to diagnose the illness are not commonly employed around the world. This work describes the coupling of microfluidic chips (MCs) to copper nanowires (CuNWs) as electrochemical detectors for the fast diagnosis of galactosemia in precious newborn urine samples. Conceptually speaking, we hypothesize that the inherent selectivity and sensitivity of CuNWs, toward galactosemia metabolites detection in connection with MC selectivity could allow the fast and simultaneous detection of the three galactosemia biomarkers, which implies the fast diagnosis of any galactosemia type in just one single analysis. Electrosynthesized CuNWs show a well-defined shape, with an average length of 6 μm and a width of 300 nm. The modified electrodes exhibited an enhanced electroactive surface area twice as high as the nonmodified ones. Very good intraelectrode repeatability with relative standard deviations (RSDs) of <8% (n = 10) and interelectrode reproducibility with RSDs of <12% (n = 5) were obtained, indicating an excellent stability of the nanoscaled electrochemical detector. Under optimum chemical (3 mM NaOH, pH 11.5), electrokinetic (separation voltage +750 V, injection +1500 V for 5 s) and electrochemical (E = +0.70 V in 3 mM NaOH, pH 11.5) conditions, galactosemia diseases were unequivocally identified, differentiating between type I, II, and III, using selected precious ill diagnosed newborn urine samples. Detection proceeded within less than 350 s, required negligible urine sample consumption, and displayed impressive signal-to-noise characteristics (ranging from 14 to 80) and micromolar limits of detection (LODs) much lower than the cutoff levels (Gal 1-P > 0.4 mM and Gal > 1.4 mM). Excellent reproducible recoveries (93%-107%, RSDs <6%) were also achieved, revealing the reliability of the approach. The significance of the newborn urine samples studied confirms the analytical potency of MC-CuNWs approach, enhancing the maturity of the microchip technology and opening new avenues for future implementation of screening applications in the field.

Reply to A. Azad et al

Reply to A. Azad et al

Chris Toumazou

Inspired by his son, an electrical engineer brings microchip technology and portability to biology.

An Hybrid Digital-Analog Sliding Mode Controller for Photovoltaic Applications

Sliding mode is a control technique that is widely employed in switching converter applications. Such a nonlinear control technique, which is commonly implemented by means of analog circuitry only, forces the switches commutations by comparing a reference signal with a combination of signals acquired on the converter itself. In this way, the control action and the switching modulation are integrated in a single functional block. In this paper the implementation of the sliding mode controller in a low cost digital device is discussed. The proposed approach profits from features offered by some digital devices, e.g., the dsPic33FJ16GS502 family from Microchip Technology Inc., in order to make the implementation simple. In the paper, the aspects related to the accuracy of the comparison between the signals required by the sliding mode controller are discussed. Moreover, the elaboration time needed to calculate the signals that drive the converter's switching devices is the subject of an in-depth analysis. The techniques proposed in the paper have been validated by means of simulations and through experimental results performed on a switching dc/dc converter for photovoltaic applications.

Bioanalysis: Heri, hodie, cras

This review, compiled for celebrating the decennial of the series of issues of Electrophoresis on bioanalysis, does not cover much heri (yesterday in Latin), since this was extensively described by my previous review Electrophoresis 2004, 25, 2111-2127, nor, for that matter cras (tomorrow in Latin) since making predictions on future scenarios is decidedly a nonscientific endeavor, but is concentrated onto hodie (today in Latin). Recent progress in the following fields is covered: SDS electrophoresis; IEF and the unraveling of the chemistry and composition of soluble carrier ampholytes; 2D maps; CE with microchip technology and some coatings of unique performance; chemistry and function of combinatorial peptide ligand libraries. This latter technique is becoming fundamental in the detection and identification of low-abundance proteins in proteome analysis.

Spatiotemporal Cattle Data—A Plea for Protocol Standardization

It was not until the end of the 1990’s that animal born satellite receivers catapulted range cattle ecology into the 21st century world of microchip technology with all of its opportunities and challenges. With the global navigation satellite system (GNSS), insight about how cattle use a landscape is being revealed from previously unknown temporal and spatial behaviors. The most common system to date for studying ungulate movement is the global positioning system (GPS). With its use has come a clarity and completeness in documenting spatial and temporal data in new and exciting ways that offer almost unlimited possibilities to better understand and manage economic and societal returns from animal dominated landscapes. However, its use on free-ranging cattle is not without challenges, some of which are yet to be optimally solved. To maximize the usefulness of GNSS data, consideration must be given to: 1) developing a standardized protocol for reporting and analyzing research that facilitates interpretation of results across different ecosystems; 2) develop optimum ranges over which to collect satellite fixes depending upon the particular behaviors of interest; and 3) concurrently develop electronic hardware and equipment platforms that are easily deployed on animals and that are light, robust, and can be worn by cattle for extended periods of time without human intervention (e.g., changing batteries). Once data are collected, appropriate geographic information system (GIS) based models should be used to produce a series of products that can be used to implement flexible management strategies, some of which may support methodologies that are yet to be commercialized and adopted into future plant-animal interface management routines.

Nanobiotechnology advanced antifouling surfaces for the continuous electrochemical monitoring of glucose in whole blood using a lab-on-a-chip

In the current work we have developed a lab-on-a-chip containing embedded amperometric sensors in four microreactors that can be addressed individually and that are coated with crystalline surface protein monolayers to provide a continuous, stable, reliable and accurate detection of blood glucose. It is envisioned that the microfluidic device will be used in a feedback loop mechanism to assess natural variations in blood glucose levels during hemodialysis to allow the individual adjustment of glucose. Reliable and accurate detection of blood glucose is accomplished by simultaneously performing (a) blood glucose measurements, (b) autocalibration routines, (c) mediator-interferences detection, and (d) background subtractions. The electrochemical detection of blood glucose variations in the absence of electrode fouling events is performed by integrating crystalline surface layer proteins (S-layer) that function as an efficient antifouling coating, a highly-oriented immobilization matrix for biomolecules and an effective molecular sieve with pore sizes of 4 to 5 nm. We demonstrate that the S-layer protein SbpA (from Lysinibacillus sphaericus CCM 2177) readily forms monomolecular lattice structures at the various microchip surfaces (e.g. glass, PDMS, platinum and gold) within 60 min, eliminating unspecific adsorption events in the presence of human serum albumin, human plasma and freshly-drawn blood samples. The highly isoporous SbpA-coating allows undisturbed diffusion of the mediator between the electrode surface, thus enabling bioelectrochemical measurements of glucose concentrations between 500 μM to 50 mM (calibration slope δI/δc of 8.7 nA mM(-1)). Final proof-of-concept implementing the four microfluidic microreactor design is demonstrated using freshly drawn blood. Accurate and drift-free assessment of blood glucose concentrations (6. 4 mM) is accomplished over 130 min at 37 °C using immobilized enzyme glucose oxidase by calculating the difference between autocalibration (10 mM glc) and background measurements. The novel combination of biologically-derived nanostructured surfaces with microchip technology constitutes a powerful new tool for multiplexed analysis of complex samples.

Single Cell Proteomics Reveals Novel Cytokine-Producing Function of Hematopoietic Stem and Progenitor Cells

AbstractAbstract 26During infection, hematopoietic stem and progenitor cells (HSPCs) are called upon to proliferate and differentiate to produce more innate and adaptive immune cells to combat infection. Traditionally, HSPCs are thought to respond to depletion of downstream hematopoietic cells during infection. More recent evidence suggests that HSPCs may respond directly to infection and pro-inflammatory cytokines. However, little is known about the direct immune response of HSPCs and the molecular signaling regulating this response upon sensing an infection. In this study, we have combined transgenic and genetic knockout mouse models with a novel single cell barcode proteomics microchip technology to tackle these questions. We show that although long-term hematopoietic stem cells (HSCs) (defined by Lineage-cKit+Sca1+CD150+CD48-) do not secrete cytokines upon toll-like receptor (TLR) stimulation, short-term HSCs and multipotent progenitor cells (MPPs) (defined by Lineage-cKit+Sca1+, referred to as LKS thereafter) can produce copious amounts of cytokines upon direct TLR-4 and TLR-2 stimulation, indicating that LKS cells can directly participate in an immune response by producing a myriad of cytokines, upon a bacterial infection. Within the population of LKS cells we detect multiple functional subsets of cells, specialized in producing myeloid-like, lymphoid-like or both types of cytokines. Moreover, we show that the cytokine production by LKS cells is regulated by the NF-κB activity, as p50-deficient LKS cells show reduced cytokine production while microRNA-146a (miR-146a)-deficient LKS cells show significantly increased cytokine production.As long-term HSCs differentiate, they start to gain effector immune function much earlier than we had originally anticipated. In light of this finding, we should start to view the stepwise differentiation scheme of HSCs, and perhaps all other stem cells, as a strategy to sequentially gain functional capacity, instead of simply losing stemness and self-renewal ability. The remarkable ability of LKS cells to produce copious amounts of cytokines in response to bacteria may provide some protective immunity during severe neutropenia and lymphopenia or in the early stage of HSC transplantation. This study further extends the functions of NF-κB to include the regulation of primitive hematopoietic stem and progenitor cells and provides direct evidence of the bacteria-responding ability of HSPCs through the TLR/NF-κB axis. The single cell barcode proteomics technology can be widely applied to study proteomics of other rare cells or heterogeneous cell population at a single cell level. Disclosures:No relevant conflicts of interest to declare.

Food analysis on microchip electrophoresis: An updated review

From 2008 to date, basically, single-cross microchip electrophoresis (ME) design has been used for food analysis with electrochemical and laser-induced fluorescence detection being the most common principles coupled. In the last 4 years, the main outlines were: (i) the exploration of new analytes such as heavy metals, nitrite, micotoxins, microorganisms, and allergens; (ii) the development of electrokinetic microfluidic (bio-) sensors into microchip format for the detection of toxins; and interestingly (iii) although sample preparation is still performed off-chip, an important increase in works dealing with complicated food samples has been clearly noticed. Although microchip technology based on electrokinetics is emerging from important fields such as authentication of foods, detection of frauds, toxics, and allergens; the marriage between micro- and nanotechnologies and total integration approaches has not reached the expected impact in the field but it is still a great promise for the development of ME of new generations for food analysis.