Disposable Microchip Research Articles

Disposable Screen-Printed Microchip Based on Nanoparticles Sensitive Membrane for Potentiometric Determination of Lead

Realization of screen-printed disposable microchip based on organic membrane sensitive layer highly responsive to lead has been demonstrated for the first time. Fabrication, potentiometric characterization and analytical application of the novel microchip have been reported. A sensitive layer comprises TiO2 nanoparticles and multiwalled carbon nanotubes “MWCNTs” composite incorporated in PVC membrane has uploaded on the plastic screen-printed microelectrode substrate surface using novel protocol. The new chip provided a linear behavior for Pb2+ ions over the lead concentration range of 1 × 10−6–1 × 10−1 mole L−1 with super Nernstian sensitivity (49 mV), relatively long life span (>4 months), and a fast response time (10 s). The advantages showed by the microchip include simple fabrication, small size, mass production, cost effectiveness, and automation and integration feasibility. The realized new microchip has been successfully utilized in the quantification of some lead (II) samples with average recovery of 101.9% and the RDS was <3.

A portable molecularly imprinted polymer-modified microchip sensor for the rapid detection of perfluorooctanoic acid.

High-performance electrochemical sensors have attracted intensive interest in real-time environmental safety monitoring, the Internet of Things, and telemedicine applications. A key limitation to field measurement of pollutant distribution is the lack of a highly sensitive and selective monitoring platform, thus severely hindering the decentralized monitoring of pollutant exposure risk. Hence, a sensor was developed in this study by using a molecularly imprinted polymer (MIP). Specifically, Cu2O@C@NiCo2O4, with a large surface area and high conductivity, was coated onto the Au electrode surface and further modified by the anodic electro-polymerization of o-phenylenediamine (o-PD) using perfluorooctanoic acid (PFOA) as the template, followed by template removal for activation, thus obtaining the Au/Cu2O@C@NiCo2O4/MIP electrode. Particularly, an effective monitoring platform derived from this sensor was designed to achieve cost-effective pollution detection. Au/Cu2O@C@NiCo2O4/MIP was employed in a disposable microchip sensor for the sensitive detection of PFOA, exhibiting an ultra-low limit of detection (LOD) of 19.46 ng L-1 in a linear range of 207-4140 ng L-1 along with satisfactory sensitivity, selectivity, and reproducibility, which reveal its great potential in the low-cost and efficient field detection of PFOA in coastal seawater. These promising results indicate a bright future for such microchip-sensor-supported PFOA tele-sensing platforms in aiding environmental safety and blue earth protection. We will persist in refining this method to enhance the sensitivity of the sensor for PFOA detection in polluted coastal areas.

Arrays of Copper Microelectrodes from Disposable Chips: Fabrication and Characterization

A simple, fast, and low-cost process to fabricate arrays of copper microelectrodes (CuMEs) based on disposable electronic microchips is described. Arrays with 8 to 20 CuMEs were characterized by energy-dispersive X-ray spectroscopy and cyclic voltammetry techniques. The closest interelectrode distance in the arrays is 358 ± 22 μm, and the minor radius ranged from 10.6 to 13.5 μm. The microchips with CuMEs were sealed in epoxy resin to fabricate the rod and flat-shaped platforms, allowing the CuMEs to be addressed separately. Glucose, hydrazine, and nitrate were used as analyte models for voltammetric and amperometric detection at CuMEs arrays, showing excellent performance in batch and flow-through cells. Glucose measurements carried out with flow injection analysis system with amperometric detection at an array of 20 CuMEs showed a wide linear range (0.020-4.0 mmol L-1), high sensitivity (734.1 μA L mmol-1 cm-2), and a limit of detection of 1.7 μmol L-1.

A confirmatory test for sperm in sexual assault samples using a microfluidic-integrated cell phone imaging system

Rapid and efficient processing of sexual assault evidence to accelerate forensic investigation and decrease casework backlogs is urgently needed. Therefore, the standardized protocols currently used in forensic laboratories can benefit from continued innovation to handle the increasing number and complexity of samples being submitted to forensic labs. To our knowledge, there is currently no available rapid and portable forensic screening technology based on a confirmatory test for sperm identification in a sexual assault kit. Here, we present a novel forensic sample screening tool, i.e., a microchip integrated with a portable cell phone imaging platform that records and processes images for further investigation and storage. The platform (i) precisely and rapidly screens swab samples (<15 min after sample preparation on-chip); (ii) selectively captures sperm from mock sexual assault samples using a novel and previously published SLeX-based surface chemistry treatment (iii) separates non-sperm contents (epithelial cells and debris in this case) out of the channel by flow prior to imaging; (iv) captures cell phone images on a portable cellphone-integrated imaging platform, (v) quantitatively differentiates sperm cells from epithelial cells, using a morphology detection code that leverages Laplacian of Gaussian and Hough gradient transform methods; (vi) is sensitive within a forensic cut-off (>95% accuracy) compared to the manual counts; (vii) provides a cost-effective and timely solution to a problem which in the past has taken a great deal of time; and (viii) handles small volumes of sample (20 μL). This integration of the cellphone imaging platform and cell recognition algorithms with disposable microchips can be a new direction toward a direct visual test to screen and differentiate sperm from epithelial cell types in forensic samples for a crime laboratory scenario. With further development, this integrated platform could assist a sexual assault nurse examiner (SANE) in a hospital or sexual assault treatment center facility to flag sperm-containing samples prior to further downstream testing.

Multifunctional microchip-based distillation apparatus I - Steam distillation for formaldehyde detection

A multifunctional microchip-based distillation apparatus for distilling and detecting formaldehyde (CH2O) in food products is developed. The presented apparatus comprises a disposable microchip, a steam supply system, and a recirculating cooling water supply. The microchip is formed on PMMA substrates by laser ablation and includes a sample zone, a flash distillation zone, a cooling zone, a condensation zone, and a collection zone. In the presented method, the CH2O sample is placed in the microchip and is vaporized by the high-throughput vapor supply and driven through the condensed zone. The condensed CH2O liquid is guided into the collection zone of the microchip. Finally, the distilled CH2O solution is determined using an AHMT spectrometry method and a paper-based RGB (red, green and blue) intensity analysis method. A distilled efficiency is as high as 98%, when a vapor stream rate is 0.4 ml/min and a distilled time is 10 min. Moreover, both detection methods show linear relationships of the corresponding CH2O concentrations. The actual sample suitability of the presented multifunctional microchip-based distillation apparatus is confirmed by analyzing the CH2O concentrations of 21 commodities.

Integrated internal standards: A sample prep-free method for better precision in microchip CE.

Point-of-care systems based on microchip capillary electrophoresis require single-use, disposable microchips prefilled with all necessary solutions so an untrained operator only needs to apply the sample and perform the analysis. While microchip fabrication can be (and has been) standardized, some manufacturing differences between microchips are unavoidable. To improve analyte precision without increasing device costs or introducing additional error sources, we recently proposed the use of integrated internal standards (ISTDs): ions added to the BGE in small concentrations which form system peaks in the electropherogram that can be used as a measurement reference. Here, we further expand this initial proof-of-principle test to study a clinically-relevant application of K ion concentrations in human blood; however, using a mock blood solution instead of real samples to avoid interference from other obstacles (e.g. cell lysis). Cs as an integrated ISTD improves repeatability of K ion migration times from 6.97% to 0.89% and the linear calibration correlation coefficient (R2 ) for K quantification from 0.851 to 0.967. Peak area repeatability improves from 11.6-13.3% to 4.75-5.04% at each K concentration above the LOQ. These results further validate the feasibility of using integrated ISTDs to improve imprecision in disposable microchip CE devices by demonstrating their application for physiological samples.

Portable semen analyzer for automated, rapid measurement of sperm motility and concentration

Semen analysis is the cornerstone of male infertility testing but many men tend to find the process of producing a sample at a lab uncomfortable and are reluctant to being tested. Our earlier work suggested that the integration of advancements in consumer electronics and microfabrication could be a powerful solution for developing a home-based male infertility screening [1]. Here, we report a versatile portable semen analyzer that can automatically perform the test and is not dependent on the smartphone camera for data acquisition, which makes it more adaptable to all of the smartphones in the market. The optical system can qualitatively classify semen samples (positive/negative) based on the World Health Organization guidelines (15 million sperm/mL and 40% motility) for seminal quality estimation [2]. When paired with any smartphone, it can also quantify sperm concentration (SC) and motility (SM). The hardware of the standalone device includes a 3D printed case, a miniaturized computer (SBC), a digital image sensor (CMOS), and DVD lenses to record videos and transmit data to a paired smartphone wirelessly. The smartphone application is designed to measure sperm concentration and motility using the transferred video from the stand-alone device. We evaluated the system using 30 semen samples and compared the smartphone-based results with manual analysis. Our custom-designed microfluidic devices enable simple, power-free semen sample handling. The loaded disposable microchips were inserted into a slot on the stand-alone device for data acquisition and analysis. The quantitative and qualitative sperm concentration and motility measurements were compared against the results obtained by manual analysis. The overall agreement between the two methods was evaluated using Bland-Altman’s absolute difference method of analysis. The accuracy of the system in normal/abnormal classification of the semen samples based on the WHO guidelines on SC and SM was calculated. Bland-Altman analysis revealed -3.3 million sperm/mL (LoA: 24.01 to -31.06) and 1.6 % (LoA: 9.6 to -6.1) mean biases for SC and SM, respectively. The accuracy of the device in classifying semen samples based on the concentration threshold of 15 million sperm/mL and the motility threshold of 40% was 100% for our tested sample set. These results suggest that the developed stand-alone device is able to measure sperm concentration and motility of an unwashed semen sample without any dependency to the smartphone camera. Such a device can be used with any smartphone model in the market without requiring any physical modification in the system. The mean processing time for transmission and analysis of a video on-phone is <10 seconds.

Improving chip-to-chip precision in disposable microchip capillary electrophoresis devices with internal standards.

To realize portable systems for routine measurements in point-of-care settings, MCE methods are required to be robust across many single-use chips. While it is well-known internal standards (ISTDs) improve run-to-run precision, a systematic investigation is necessary to determine the significance of chip-to-chip imprecision in MCE and how ISTDs account for it. This paper addresses this question by exploring the reproducibility of Na quantification across six basic, in-house fabricated microchips. A dataset of 900 electrophoerograms was collected from analyzing five concentrations of NaCl with two ISTDs (CsCl and LiCl). While both improved the peak area reproducibility, the Na/Cs ratio was superior to the Na/Li ratio (improving the RSD by a factor of 2-4, depending on the Na concentration). We attribute this to the significant variation in microchannel surface properties, which was accounted for by cesium but not lithium. Microchip dimension and detector variations were only a few percent, and could be improved through commercial fabrication over in-house made microchips. These results demonstrate that ISTDs not only correct for intrachip imprecision, but are also a viable means to correct for chip-to-chip imprecision inherent in disposable, point-of-care MCE devices. However, as expected, the internal standard must be carefully chosen.

Generation of Complex, Dynamic Temperature Gradients in a Disposable Microchip

We present a temperature gradient (TG) generation method using acoustothermal heating of polydimethylsiloxane (PDMS) by surface acoustic wave (SAW) microfluidic system. Independent and precise control of each SAW from a slanted interdigital transducer (IDT) enables a one-dimensional (1D) spatiotemporal control of temperature in a PDMS microchip. Various TGs were created in gas and liquid. The system creates switchable, dynamic and complex TGs in a disposable microchip.

Microchip electrophoresis for fast and interference-free determination of trace amounts of glyphosate and glufosinate residues in agricultural products.

Fast screening of herbicide residues is becoming important to ensure food safety, but traditional chromatographic methods may not be suitable for rapid on-site analysis of samples with complicated matrices. Here, we describe a method for rapid and sensitive determination of glyphosate (GLYP) and glufosinate (GLUF) residues in agricultural products by electrophoresis on disposable microchips with laser-induced fluorescence detection. With this method, quantitative analysis of trace amounts of GLYP and GLUF can be achieved with relatively simple sample preparation.

Development of On-Chip Multi-Imaging Flow Cytometry for Identification of Imaging Biomarkers of Clustered Circulating Tumor Cells

An on-chip multi-imaging flow cytometry system has been developed to obtain morphometric parameters of cell clusters such as cell number, perimeter, total cross-sectional area, number of nuclei and size of clusters as “imaging biomarkers”, with simultaneous acquisition and analysis of both bright-field (BF) and fluorescent (FL) images at 200 frames per second (fps); by using this system, we examined the effectiveness of using imaging biomarkers for the identification of clustered circulating tumor cells (CTCs). Sample blood of rats in which a prostate cancer cell line (MAT-LyLu) had been pre-implanted was applied to a microchannel on a disposable microchip after staining the nuclei using fluorescent dye for their visualization, and the acquired images were measured and compared with those of healthy rats. In terms of the results, clustered cells having (1) cell area larger than 200 µm2 and (2) nucleus area larger than 90 µm2 were specifically observed in cancer cell-implanted blood, but were not observed in healthy rats. In addition, (3) clusters having more than 3 nuclei were specific for cancer-implanted blood and (4) a ratio between the actual perimeter and the perimeter calculated from the obtained area, which reflects a shape distorted from ideal roundness, of less than 0.90 was specific for all clusters having more than 3 nuclei and was also specific for cancer-implanted blood. The collected clusters larger than 300 µm2 were examined by quantitative gene copy number assay, and were identified as being CTCs. These results indicate the usefulness of the imaging biomarkers for characterizing clusters, and all of the four examined imaging biomarkers—cluster area, nuclei area, nuclei number, and ratio of perimeter—can identify clustered CTCs in blood with the same level of preciseness using multi-imaging cytometry.

Microwave-assisted derivatization for fast and efficient analysis of saccharides on disposable microchips

A domestic microwave oven was used to achieve rapid derivatization of saccharides for their microchip electrophoresis analysis.

Development of a real-time microchip PCR system for portable plant disease diagnosis.

Rapid and accurate detection of plant pathogens in the field is crucial to prevent the proliferation of infected crops. Polymerase chain reaction (PCR) process is the most reliable and accepted method for plant pathogen diagnosis, however current conventional PCR machines are not portable and require additional post-processing steps to detect the amplified DNA (amplicon) of pathogens. Real-time PCR can directly quantify the amplicon during the DNA amplification without the need for post processing, thus more suitable for field operations, however still takes time and require large instruments that are costly and not portable. Microchip PCR systems have emerged in the past decade to miniaturize conventional PCR systems and to reduce operation time and cost. Real-time microchip PCR systems have also emerged, but unfortunately all reported portable real-time microchip PCR systems require various auxiliary instruments. Here we present a stand-alone real-time microchip PCR system composed of a PCR reaction chamber microchip with integrated thin-film heater, a compact fluorescence detector to detect amplified DNA, a microcontroller to control the entire thermocycling operation with data acquisition capability, and a battery. The entire system is 25×16×8 cm3 in size and 843 g in weight. The disposable microchip requires only 8-µl sample volume and a single PCR run consumes 110 mAh of power. A DNA extraction protocol, notably without the use of liquid nitrogen, chemicals, and other large lab equipment, was developed for field operations. The developed real-time microchip PCR system and the DNA extraction protocol were used to successfully detect six different fungal and bacterial plant pathogens with 100% success rate to a detection limit of 5 ng/8 µl sample.

Point of care creatinine measurement for diagnosis of renal disease using a disposable microchip

A point-of-care device for the determination of elevated creatinine levels in blood is reported. This device potentially offers a new and simple clinical regime for the determination of creatinine that will give huge time savings and removal of several steps of determination. The test employs a disposable prefilled microchip and the handheld Medimate Multireader®. By optimizing the analytical conditions it was found that the LOD of the proposed method was 87 μM creatinine, close to the normal human serum levels that are in the range of 60 to 100μM. A statistical analysis of the residual shows a normal distribution, indicating the absence of systematic errors in the proposed method. The test can be used to distinguish patients with renal insufficiency (creatinine levels >100μM) from healthy persons. Long-term monitoring could furthermore distinguish between acute renal failure and chronic kidney disease by the rate of creatinine concentration rise.

A Sensitive Microchip‐Based Immunosensor for Electrochemical Detection of Low‐Level Biomarker S100B

AbstractA sensitive microchip‐based electrochemical immunosensor has been developed for quantification of low level S100B, which is an important biomarker of neurological diseases. Poly(ethyleneimine) (PEI), containing NH2 groups, was firstly modified on the surface of the poly(methyl methacrylate) (PMMA) microchannel to capture monoclonal anti‐S100B. Afterward, S100B and second polyclonal anti‐S100 were sequentially coupled by antigen‐antibody specific interaction, and the alkaline phosphatase conjugates (ALP‐conjugates) were bound to the second antibodies in a PMMA microchannnel. Using differential pulse voltammetry (DPV), the immunosensor achieved the detection limit of 0.1 pg mL−1 with the linear concentration range of 0.1 pg mL−1–100 pg mL−1. The results indicate that the proposed strategy not only has high specificity and sensitivity, but also resists nonspecific protein adsorption. The disposable microchip immunosensor shows good promise for clinical screening of important biomarkers and convenient point‐of‐care diagnostics.

Design of disposable DNA biosensor microchip with amperometric detection featuring PCB substrate

Electrochemical detection (ECD) is one of the best means for designing a Lab-on-a-chip system especially for electroactive analyte’s separation and detection applications like biomedical and in vitro diagnostic. Producing disposable biosensors is the key element in the recent research. Nevertheless, cost of the fabrication is the main obstacle against producing affordable and disposable microchips. In this paper, a workable DNA separation and detection microchip based on printed circuit board (PCB) substrates is presented. The new design will help producing lowcost Lab-on-a-chip devices for DNA or other electroactive diagnostic purposes. Experiments performed using standard DNA ladder produced results which are comparable with the conventional gel electrophoresis. With sensitivity consistently higher than 100 nA and separation time of approximately 30 min, this new design is an ideal tool for real DNA analysis.

Rapid patterning of ‘tunable’ hydrophobic valves on disposable microchips by laser printer lithography

We recently defined a method for fabricating multilayer microdevices using poly(ethylene terephthalate) transparency film and printer toner, and showed these could be successfully applied to DNA extraction and amplification (Duarte et al., Anal. Chem. 2011, 83, 5182-5189). Here, we advance the functionality of these microdevices with flow control enabled by hydrophobic valves patterned using laser printer lithography. Laser printer patterning of toner within the microchannel induces a dramatic change in surface hydrophobicity (change in contact angle of DI water from 51° to 111°) with good reproducibility. Moreover, the hydrophobicity of the surface can be controlled by altering the density of the patterned toner via varying the gray-scale setting on the laser printer, which consequently tunes the valve's burst pressure. Toner density provided a larger burst pressure bandwidth (158 ± 18 Pa to 573 ± 16 Pa) than could be achieved by varying channel geometry (492 ± 18 Pa to 573 ± 16 Pa). Finally, we used a series of tuned toner valves (with varied gray-scale) for passive valve-based fluidic transfer in a predictable manner through the architecture of a rotating PeT microdevice. While an elementary demonstration, this presents the possibility for simplistic and cost-effective microdevices with valved fluid flow control to be fabricated using nothing more than a laser printer, a laser cutter and a laminator.

Enumeration of CD4+ T-Cells Using a Portable Microchip Count Platform in Tanzanian HIV-Infected Patients

BackgroundCD4+ T-lymphocyte count (CD4 count) is a standard method used to monitor HIV-infected patients during anti-retroviral therapy (ART). The World Health Organization (WHO) has pointed out or recommended that a handheld, point-of-care, reliable, and affordable CD4 count platform is urgently needed in resource-scarce settings.MethodsHIV-infected patient blood samples were tested at the point-of-care using a portable and label-free microchip CD4 count platform that we have developed. A total of 130 HIV-infected patient samples were collected that included 16 de-identified left over blood samples from Brigham and Women's Hospital (BWH), and 114 left over samples from Muhimbili University of Health and Allied Sciences (MUHAS) enrolled in the HIV and AIDS care and treatment centers in the City of Dar es Salaam, Tanzania. The two data groups from BWH and MUHAS were analyzed and compared to the commonly accepted CD4 count reference method (FACSCalibur system).ResultsThe portable, battery operated and microscope-free microchip platform developed in our laboratory (BWH) showed significant correlation in CD4 counts compared with FACSCalibur system both at BWH (r = 0.94, p<0.01) and MUHAS (r = 0.49, p<0.01), which was supported by the Bland-Altman methods comparison analysis. The device rapidly produced CD4 count within 10 minutes using an in-house developed automated cell counting program.ConclusionsWe obtained CD4 counts of HIV-infected patients using a portable platform which is an inexpensive (<$1 material cost) and disposable microchip that uses whole blood sample (<10 µl) without any pre-processing. The system operates without the need for antibody-based fluorescent labeling and expensive fluorescent illumination and microscope setup. This portable CD4 count platform displays agreement with the FACSCalibur results and has the potential to expand access to HIV and AIDS monitoring using fingerprick volume of whole blood and helping people who suffer from HIV and AIDS in resource-limited settings.

A CCD-based fluorescence imaging system for real-time loop-mediated isothermal amplification-based rapid and sensitive detection of waterborne pathogens on microchips

Rapid, sensitive, and low-cost pathogen diagnostic systems are needed for early disease diagnosis and treatment, especially in resource-limited settings. This study reports a low-cost charge-coupled device (CCD)-based fluorescence imaging system for rapid detection of waterborne pathogens by isothermal gene amplification in disposable microchips. Fluorescence imaging capability of this monochromatic CCD camera is evaluated by optimizing the gain, offset, and exposure time. This imaging system is validated for 12 virulence genes of major waterborne pathogens on cyclic olefin polymer (COP) microchips, using SYTO-82 dye and real time fluorescence loop-mediated isothermal amplification referred here as microRT(f)-LAMP. Signal-to-noise ratio (SNR) and threshold time (Tt) of microRT(f)-LAMP assays are compared with those from a commercial real-time polymerase chain reaction (PCR) instrument. Applying a CCD exposure of 5 s to 10(5) starting DNA copies of microRT(f)-LAMP assays increases the SNR by 8-fold and reduces the Tt by 9.8 min in comparison to a commercial real-time PCR instrument. Additionally, single copy level sensitivity for Campylobacter jejuni 0414 gene is obtained for microRT(f)-LAMP with a Tt of 19 min, which is half the time of the commercial real-time PCR instrument. Due to the control over the exposure time and the wide field imaging capability of CCD, this low-cost fluorescence imaging system has the potential for rapid and parallel detection of pathogenic microorganisms in high throughput microfluidic chips.

Gold Magnetic Particle Based Immunoassay on a Disposable Microchips

Microchips appear to offer important opportunities for modern research. Microchips based on gold magnetic nanoparticles make manipulation of biomolecules more conveniently, and has the advantage of deducing reaction time. In this study, we first achieved microchips design and fabrication through CO2 laser ablation for immunoassay, and then conjugated mouse IgG to gold magnetic nanoparticles serve as a target. Finally, HRP labeled goat anti mouse IgG binding assay and substrate reaction were performed with disposable microchips and other devices. The results indicated that the reactions carried on successfully and magnetic particles moved well in chips.