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1.
Anal Bioanal Chem ; 414(8): 2607-2618, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35091761

RESUMO

The lateral flow assay (LFA) is one of the most popular technologies on the point-of-care diagnostics market due to its low cost and ease of use, with applications ranging from pregnancy to environmental toxins to infectious disease. While the use of these tests is relatively straightforward, significant development time and effort are required to create tests that are both sensitive and specific. Workflows to guide the LFA development process exist but moving from target selection to an LFA that is ready for field testing can be labor intensive, resource heavy, and time consuming. To reduce the cost and the duration of the LFA development process, we introduce a novel development platform centered on the flexibility, speed, and throughput of an automated robotic liquid handling system. The system comprises LFA-specific hardware and software that enable large optimization experiments with discrete and continuous variables such as antibody pair selection or reagent concentration. Initial validation of the platform was demonstrated during development of a malaria LFA but was readily expanded to encompass development of SARS-CoV-2 and Mycobacterium tuberculosis LFAs. The validity of the platform, where optimization experiments are run directly on LFAs rather than in solution, was based on a direct comparison between the robotic system and a more traditional ELISA-like method. By minimizing hands-on time, maximizing experiment size, and enabling improved reproducibility, the robotic system improved the quality and quantity of LFA assay development efforts.


Assuntos
COVID-19/diagnóstico , Imunoensaio/instrumentação , Malária/diagnóstico , Testes Imediatos , Tuberculose/diagnóstico , Teste Sorológico para COVID-19/economia , Teste Sorológico para COVID-19/instrumentação , Desenho de Equipamento , Humanos , Imunoensaio/economia , Mycobacterium tuberculosis/isolamento & purificação , Plasmodium/isolamento & purificação , Testes Imediatos/economia , Reprodutibilidade dos Testes , SARS-CoV-2/isolamento & purificação , Sensibilidade e Especificidade , Fatores de Tempo
2.
Anal Chem ; 90(11): 6643-6650, 2018 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-29683653

RESUMO

Lateral flow assays (LFAs) are widely used for yes/no detection of analytes, but they are not well-suited for quantification. We show that the sensitivity of the test line in a lateral flow assay can be tuned to appear at a specific sample concentration by varying the density of capture molecules at the test line and that when test lines tuned for different responses are combined into a single test strip, lines appear at specific thresholds of sample concentration. We also developed a model based on mass-action kinetics that accurately described test line signal and shape over a wide matrix of capture molecules and sample concentrations in single-line strips. The model was used to design a three-line test strip with lines designed to appear at logarithmically spaced sample concentrations, and the experiments showed a remarkable match to predictions. The response of this "graded ladder bar" format is due to the effect of test line concentration on capture efficiency at each test line, not on sample depletion effects, and the effect is maintained whether a system is under kinetic or equilibrium control. These features enable design of nonlinear responses (logarithmic here) and suggest robustness for different systems. Thus, the graded ladder bar format could be a useful tool for applications requiring quantification of sample concentrations over a wide dynamic range.

3.
Environ Sci Technol ; 52(6): 3567-3573, 2018 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-29470061

RESUMO

Metal contamination of natural and drinking water systems poses hazards to public and environmental health. Quantifying metal concentrations in water typically requires sample collection in the field followed by expensive laboratory analysis that can take days to weeks to obtain results. The objective of this work was to develop a low-cost, field-deployable method to quantify trace levels of copper in drinking water by coupling solid-phase extraction/preconcentration with a microfluidic paper-based analytical device. This method has the advantages of being hand-powered (instrument-free) and using a simple "read by eye" quantification motif (based on color distance). Tap water samples collected across Fort Collins, CO, were tested with this method and validated against ICP-MS. We demonstrate the ability to quantify the copper content of tap water within 30% of a reference technique at levels ranging from 20 to 500 000 ppb. The application of this technology, which should be sufficient as a rapid screening tool, can lead to faster, more cost-effective detection of soluble metals in water systems.


Assuntos
Água Potável , Poluentes Químicos da Água , Cobre , Água Doce , Extração em Fase Sólida
4.
Analyst ; 141(6): 1874-87, 2016 Mar 21.
Artigo em Inglês | MEDLINE | ID: mdl-26901771

RESUMO

The field of paper-based microfluidics has experienced rapid growth over the past decade. Microfluidic paper-based analytical devices (µPADs), originally developed for point-of-care medical diagnostics in resource-limited settings, are now being applied in new areas, such as environmental analyses. Low-cost paper sensors show great promise for on-site environmental analysis; the theme of ongoing research complements existing instrumental techniques by providing high spatial and temporal resolution for environmental monitoring. This review highlights recent applications of µPADs for environmental analysis along with technical advances that may enable µPADs to be more widely implemented in field testing.


Assuntos
Técnicas de Química Analítica/métodos , Meio Ambiente , Papel , Animais , Técnicas de Química Analítica/instrumentação , Humanos
5.
Ann Occup Hyg ; 58(4): 413-23, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24515892

RESUMO

Metals in particulate matter (PM) are considered a driving factor for many pathologies. Despite the hazards associated with particulate metals, personal exposures for at-risk workers are rarely assessed due to the cost and effort associated with monitoring. As a result, routine exposure assessments are performed for only a small fraction of the exposed workforce. The objective of this research was to evaluate a relatively new technology, microfluidic paper-based analytical devices (µPADs), for measuring the metals content in welding fumes. Fumes from three common welding techniques (shielded metal arc, metal inert gas, and tungsten inert gas welding) were sampled in two welding shops. Concentrations of acid-extractable Fe, Cu, Ni, and Cr were measured and independently verified using inductively coupled plasma-optical emission spectroscopy (ICP-OES). Results from the µPAD sensors agreed well with ICP-OES analysis; the two methods gave statistically similar results in >80% of the samples analyzed. Analytical costs for the µPAD technique were ~50 times lower than market-rate costs with ICP-OES. Further, the µPAD method was capable of providing same-day results (as opposed several weeks for ICP laboratory analysis). Results of this work suggest that µPAD sensors are a viable, yet inexpensive alternative to traditional analytic methods for transition metals in welding fume PM. These sensors have potential to enable substantially higher levels of hazard surveillance for a given resource cost, especially in resource-limited environments.


Assuntos
Poluentes Ocupacionais do Ar/análise , Microfluídica/métodos , Exposição Ocupacional/análise , Soldagem , Poluentes Ocupacionais do Ar/economia , Cromo/análise , Monitoramento Ambiental/instrumentação , Humanos , Microfluídica/economia , Níquel/análise , Material Particulado/análise , Medição de Risco , Aço Inoxidável/química
6.
Am J Trop Med Hyg ; 106(3): 850-852, 2022 01 17.
Artigo em Inglês | MEDLINE | ID: mdl-35026727

RESUMO

Rapid diagnostic tests (RDTs) for Plasmodium falciparum commonly detect histidine-rich protein 2 (HRP-2), but HRP-2 deletions are increasingly recognized. We evaluated a prototype test detecting parasite lactate dehydrogenase (pLDH) and compared it to commercially available RDTs at a health facility in Uganda, using quantitative polymerase chain reaction as a gold standard. The prototype pLDH test had a high sensitivity for infections with at least 100 parasites/µL (98%), comparable to HRP-2, and greater than an existing pLDH RDT (89%). Specificity for the prototype test was 99.5%, which is greater than the HRP-2 tests (93-95%). Therefore, the prototype pLDH test may be an attractive alternative malaria diagnostic.


Assuntos
Malária Falciparum , Malária , Antígenos de Protozoários/análise , Testes Diagnósticos de Rotina , Humanos , L-Lactato Desidrogenase/análise , Malária/diagnóstico , Malária Falciparum/diagnóstico , Malária Falciparum/parasitologia , Microscopia , Plasmodium falciparum , Reação em Cadeia da Polimerase , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Sensibilidade e Especificidade , Uganda
7.
PLoS One ; 16(8): e0256352, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34403456

RESUMO

Rapid tests for SARS-COV-2 infection are important tools for pandemic control, but current rapid tests are based on proprietary designs and reagents. We report clinical validation results of an open-access lateral flow assay (OA-LFA) design using commercially available materials and reagents, along with RT-qPCR and commercially available comparators (BinaxNOW® and Sofia®). Adult patients with suspected COVID-19 based on clinical signs and symptoms, and with symptoms ≤7 days duration, underwent anterior nares (AN) sampling for the OA-LFA, Sofia®, BinaxNOW ™, and RT-qPCR, along with nasopharyngeal (NP) RT-qPCR. Results indicate a positive predictive agreement with NP sampling as 69% (60% -78%) OA-LFA, 74% (64% - 82%) Sofia®, and 82% (73% - 88%) BinaxNOW™. The implication for these results is that we provide an open-access LFA design that meets the minimum WHO target product profile for a rapid test, that virtually any diagnostic manufacturer could produce.


Assuntos
Antígenos Virais/análise , COVID-19/diagnóstico , Imunoensaio , SARS-CoV-2/metabolismo , Área Sob a Curva , COVID-19/virologia , Humanos , Nasofaringe/virologia , Sistemas Automatizados de Assistência Junto ao Leito , RNA Viral/análise , RNA Viral/metabolismo , Curva ROC , Reação em Cadeia da Polimerase em Tempo Real , SARS-CoV-2/isolamento & purificação , Sensibilidade e Especificidade
8.
ACS Omega ; 6(39): 25116-25123, 2021 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-34608447

RESUMO

The global COVID-19 pandemic has created an urgent demand for large numbers of inexpensive, accurate, rapid, point-of-care diagnostic tests. Analyte-based assays are suitably rapid and inexpensive and can be rapidly mass-produced, but for sufficiently accurate performance, they require highly optimized antibodies and assay conditions. We used an automated liquid handling system, customized to handle arrays of lateral flow (immuno)assays (LFAs) in a high-throughput screen, to identify anti-nucleocapsid antibodies that will perform optimally in an LFA. We tested 1021 anti-nucleocapsid antibody pairs as LFA capture and detection reagents with the goal of highlighting pairs that have the greatest affinity for the nucleocapsid protein of SARS-CoV-2 within the LFA format. In contrast to traditional antibody screening methods (e.g., ELISA, bio-layer interferometry), the method described here integrates real-time reaction kinetics with transport in, and immobilization directly onto, nitrocellulose. We have identified several candidate antibody pairs that are suitable for further development of an LFA for SARS-CoV-2.

9.
ACS Omega ; 6(31): 20139-20148, 2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34373846

RESUMO

Severe acute respiratory coronavirus-2 (SARS-CoV-2) is a novel viral pathogen and therefore a challenge to accurately diagnose infection. Asymptomatic cases are common and so it is difficult to accurately identify infected cases to support surveillance and case detection. Diagnostic test developers are working to meet the global demand for accurate and rapid diagnostic tests to support disease management. However, the focus of many of these has been on molecular diagnostic tests, and more recently serologic tests, for use in primarily high-income countries. Low- and middle-income countries typically have very limited access to molecular diagnostic testing due to fewer resources. Serologic testing is an inappropriate surrogate as the early stages of infection are not detected and misdiagnosis will promote continued transmission. Detection of infection via direct antigen testing may allow for earlier diagnosis provided such a method is sensitive. Leading SARS-CoV-2 biomarkers include spike protein, nucleocapsid protein, envelope protein, and membrane protein. This research focuses on antibodies to SARS-CoV-2 spike protein due to the number of monoclonal antibodies that have been developed for therapeutic research but also have potential diagnostic value. In this study, we assessed the performance of antibodies to the spike glycoprotein, acquired from both commercial and private groups in multiplexed liquid immunoassays, with concurrent testing via a half-strip lateral flow assays (LFA) to indicate antibodies with potential in LFA development. These processes allow for the selection of pairs of high-affinity antispike antibodies that are suitable for liquid immunoassays and LFA, some of which with sensitivity into the low picogram range with the liquid immunoassay formats with no cross-reactivity to other coronavirus S antigens. Discrepancies in optimal ranking were observed with the top pairs used in the liquid and LFA formats. These findings can support the development of SARS-CoV-2 LFAs and diagnostic tools.

11.
Anal Chim Acta ; 915: 64-73, 2016 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-26995641

RESUMO

Measurement variance is a crucial aspect of quantitative chemical analysis. Variance directly affects important analytical figures of merit, including detection limit, quantitation limit, and confidence intervals. Most reported analyses for emerging analytical techniques implicitly assume constant variance (homoskedasticity) by using unweighted regression calibrations. Despite the assumption of constant variance, it is known that most instruments exhibit heteroskedasticity, where variance changes with signal intensity. Ignoring nonconstant variance results in suboptimal calibrations, invalid uncertainty estimates, and incorrect detection limits. Three techniques where homoskedasticity is often assumed were covered in this work to evaluate if heteroskedasticity had a significant quantitative impact-naked-eye, distance-based detection using paper-based analytical devices (PADs), cathodic stripping voltammetry (CSV) with disposable carbon-ink electrode devices, and microchip electrophoresis (MCE) with conductivity detection. Despite these techniques representing a wide range of chemistries and precision, heteroskedastic behavior was confirmed for each. The general variance forms were analyzed, and recommendations for accounting for nonconstant variance discussed. Monte Carlo simulations of instrument responses were performed to quantify the benefits of weighted regression, and the sensitivity to uncertainty in the variance function was tested. Results show that heteroskedasticity should be considered during development of new techniques; even moderate uncertainty (30%) in the variance function still results in weighted regression outperforming unweighted regressions. We recommend utilizing the power model of variance because it is easy to apply, requires little additional experimentation, and produces higher-precision results and more reliable uncertainty estimates than assuming homoskedasticity.

12.
Lab Chip ; 15(13): 2808-18, 2015 Jul 07.
Artigo em Inglês | MEDLINE | ID: mdl-26009988

RESUMO

Exposure to metal-containing aerosols has been linked with adverse health outcomes for almost every organ in the human body. Commercially available techniques for quantifying particulate metals are time-intensive, laborious, and expensive; often sample analysis exceeds $100. We report a simple technique, based upon a distance-based detection motif, for quantifying metal concentrations of Ni, Cu, and Fe in airborne particulate matter using microfluidic paper-based analytical devices. Paper substrates are used to create sensors that are self-contained, self-timing, and require only a drop of sample for operation. Unlike other colorimetric approaches in paper microfluidics that rely on optical instrumentation for analysis, with distance-based detection, analyte is quantified visually based on the distance of a colorimetric reaction, similar to reading temperature on a thermometer. To demonstrate the effectiveness of this approach, Ni, Cu, and Fe were measured individually in single-channel devices; detection limits as low as 0.1, 0.1, and 0.05 µg were reported for Ni, Cu, and Fe. Multiplexed analysis of all three metals was achieved with detection limits of 1, 5, and 1 µg for Ni, Cu, and Fe. We also extended the dynamic range for multi-analyte detection by printing concentration gradients of colorimetric reagents using an off-the-shelf inkjet printer. Analyte selectivity was demonstrated for common interferences. To demonstrate utility of the method, Ni, Cu, and Fe were measured from samples of certified welding fume; levels measured with paper sensors matched known values determined gravimetrically.


Assuntos
Colorimetria , Cobre/análise , Ferro/análise , Níquel/análise , Papel , Corantes/química , Microfluídica/instrumentação
13.
Anal Chim Acta ; 874: 40-8, 2015 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-25910444

RESUMO

This work describes the development of an electrochemical sensor for simultaneous detection of Zn(II), Cd(II), and Pb(II) using a graphene-polyaniline (G/PANI) nanocomposite electrode prepared by reverse-phase polymerization in the presence of polyvinylpyrrolidone (PVP). Two substrate materials (plastic film and filter paper) and two nanocomposite deposition methods (drop-casting and electrospraying) were investigated. Square-wave anodic stripping voltammetry currents were higher for plastic vs. paper substrates. Performance of the G/PANI nanocomposites was characterized by scanning electron microscopy (SEM) and cyclic voltammetry. The G/PANI-modified electrode exhibited high electrochemical conductivity, producing a three-fold increase in anodic peak current (vs. the unmodified electrode). The G/PANI-modified electrode also showed evidence of increased surface area under SEM. Square-wave anodic stripping voltammetry was used to measure Zn(II), Cd(II), and Pb(II) in the presence of Bi(III). A linear working range of 1-300 µg L(-1) was established between anodic current and metal ion concentration with detection limits (S/N=3) of 1.0 µg L(-1) for Zn(II), and 0.1 µg L(-1) for both Cd(II) and Pb(II). The G/PANI-modified electrode allowed selective determination of the target metals in the presence of common metal interferences including Mn(II), Cu(II), Fe(III), Fe(II), Co(III), and Ni(II). Repeat assays on the same device demonstrated good reproducibility (%RSD<11) over 10 serial runs. Finally, this system was utilized for determining Zn(II), Cd(II), and Pb(II) in human serum using the standard addition method.


Assuntos
Compostos de Anilina/química , Cádmio/sangue , Técnicas Eletroquímicas/instrumentação , Grafite/química , Chumbo/sangue , Zinco/sangue , Cádmio/análise , Desenho de Equipamento , Humanos , Chumbo/análise , Limite de Detecção , Nanocompostos/química , Nanocompostos/ultraestrutura , Zinco/análise
14.
Anal Methods ; 6(20): 8180-8186, 2014 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-25267929

RESUMO

Development and characterization of a simple microfluidic electrochemical flow cell that can be coupled with HPLC to enable dual absorbance/electrochemical detection is described. Coupling absorbance and electrochemical detection increases the information that can be gathered from a single injection, but a second (typically expensive) detection system is required. Here, an inexpensive, customizable microfluidic electrochemical detector is coupled in series with a commercial HPLC/UV system. The microfluidic device is made from poly(dimethylsiloxane) and contains carbon paste electrodes. To demonstrate the utility of this dual-detection system, the reaction products of the radical scavenging agent salicylic acid and hydroxyl radical generated by Fenton chemistry were analyzed. The dual-detection system was used to quantify 2,5-dihydroxybenzoic acid, 2,3-dihydroxybenzoic acid, and catechol produced by the addition of H2O2 to filter samples of welding fumes. Measurement recovery was high, with percent recoveries between 97-102%, 92-103%, and 95-103% for 2,5-dihydroxybenzoic acid, 2,3-dihydroxybenzoic acid, and catechol, respectively, for control samples. The methods described in this work are simple, reliable, and can inexpensively couple electrochemical detection to HPLC-UV systems.

15.
Lab Chip ; 13(12): 2397-404, 2013 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-23657627

RESUMO

Paper-based analytical devices (PADs) represent a growing class of elegant, yet inexpensive chemical sensor technologies designed for point-of-use applications. Most PADs, however, still utilize some form of instrumentation such as a camera for quantitative detection. We describe here a simple technique to render PAD measurements more quantitative and straightforward using the distance of colour development as a detection motif. The so-called distance-based detection enables PAD chemistries that are more portable and less resource intensive compared to classical approaches that rely on the use of peripheral equipment for quantitative measurement. We demonstrate the utility and broad applicability of this technique with measurements of glucose, nickel, and glutathione using three different detection chemistries: enzymatic reactions, metal complexation, and nanoparticle aggregation, respectively. The results show excellent quantitative agreement with certified standards in complex sample matrices. This work provides the first demonstration of distance-based PAD detection with broad application as a class of new, inexpensive sensor technologies designed for point-of-use applications.


Assuntos
Técnicas Analíticas Microfluídicas/métodos , Papel , 3,3'-Diaminobenzidina/química , Glicemia/análise , Glucose/análise , Glucose Oxidase/metabolismo , Glutationa/análise , Glutationa/sangue , Humanos , Nanopartículas Metálicas/química , Técnicas Analíticas Microfluídicas/instrumentação , Níquel/análise , Oximas/química , Peroxidase/metabolismo , Prata/química
16.
Anal Chim Acta ; 800: 50-5, 2013 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-24120167

RESUMO

Occupational exposure to Cr is concerning because of its myriad of health effects. Assessing chromium exposure is also cost and resource intensive because the analysis typically uses sophisticated instrumental techniques like inductively coupled plasma-mass spectrometry (ICP-MS). Here, we report a novel, simple, inexpensive microfluidic paper-based analytical device (µPAD) for measuring total Cr in airborne particulate matter. In the µPAD, tetravalent cerium (Ce(IV)) was used in a pretreatment zone to oxidize all soluble Cr to Cr(VI). After elution to the detection zone, Cr(VI) reacts with 1,5-diphenylcarbazide (1,5-DPC) forming 1,5-diphenylcarbazone (DPCO) and Cr(III). The resulting Cr(III) forms a distinct purple colored complex with the DPCO. As proof-of-principle, particulate matter (PM) collected on a sample filter was analyzed with the µPAD to quantify the mass of total Cr. A log-linear working range (0.23-3.75 µg; r(2)=0.998) between Cr and color intensity was obtained with a detection limit of 0.12 µg. For validation, a certified reference containing multiple competing metals was analyzed. Quantitative agreement was obtained between known Cr levels in the sample and the Cr measured using the µPAD.


Assuntos
Cromo/análise , Colorimetria/instrumentação , Técnicas Analíticas Microfluídicas/instrumentação , Papel , Material Particulado/química , Cério/química , Difenilcarbazida/química , Concentração de Íons de Hidrogênio , Micro-Ondas , Ácido Nítrico/química , Oxirredução , Polietilenos/química , Compostos de Amônio Quaternário/química
17.
Biomicrofluidics ; 4(4): 44105, 2010 Nov 02.
Artigo em Inglês | MEDLINE | ID: mdl-21124751

RESUMO

Control of the 3D microenvironment for cultured cells is essential for understanding the complex relationships that biomolecular concentration gradients have on cellular growth, regeneration, and differentiation. This paper reports a microfluidic device for delivering gradients of soluble molecules to cells in an open reservoir without exposing the cells to flow. The cells are cultured on a polyester membrane that shields them from the flow that delivers the gradient. A novel "lid" design is implemented which prevents leakage from around the membrane without requiring sealing agents or adhesives. Once layers are molded, device fabrication can be performed within minutes while at room temperature. Surface gradients were characterized with epifluorescence microscopy; image analysis verified that sharp gradients (∼33 µm wide) can be reproducibly generated. We show that heterogeneous laminar flow patterns of Orange and Green Cell Tracker (CT) applied beneath the membrane can be localized to cells cultured on the other side; concentration profile scans show the extent of CT diffusion parallel to the membrane's surface to be 10-20 µm. Our device is ideal for conventional cell culture because the cell culture surface is readily accessible to physical manipulation (e.g., micropipette access), the cell culture medium is in direct contact with the incubator atmosphere (i.e., no special protocols for ensuring proper equilibration of gas concentrations are required), and the cells are not subjected to flow-induced shear forces, which are advantageous attributes not commonly found in closed-channel microfluidic designs.

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