The spike glycoprotein of SARS-CoV-2 is a highly conserved surface protein and as such may represent a good target for immunoassay detection. We screened a variety of antibodies that were reactive to the S glycoprotein in a highly sensitive liquid immunoassay format and also on paper-based or lateral flow assay (LFA) to assess their analytical performance. Our findings included significant variation in performance when using different sources of S antigen. We identified several antibody pairs that had an LOD of below 10 pg/mL in the liquid immunoassay format with the lowest being 3 pg/mL. The antibodies were highly specific to SARS-Cov-2 based on cross reactivity screening with other human CoVs. The LFA screening found some different optimal antibody pairs from the pool of candidate antibodies used but a several antibodies were observed to have high performance with either immunoassay format.
<p></p><p>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 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 immunoassay (LFA) tests 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 unique epitopes of 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.</p><p></p>
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.
A need exists to develop simple, low-cost methods for aerosol speciation and air quality measurement to support both medium-to-large-scale epidemiology and citizen-based science. Our group has developed alternative approaches for aerosol speciation using microfluidic technologies designed to manipulate teardrop sample volumes. Traditional and paper-based microfluidics are poised to overcome many issues associated with chemical analysis of particulate matter (PM), namely: cost, ease-of-use, and sensitivity. Results can be read using either the naked eye or simple, low-cost instrumentation (i.e., a handheld or mobile device). Such assays cost only a few dollars and can be conducted by virtually anyone after simple training. We have developed microfluidic assays to measure reactive oxygen species and metals (Fe, Ni, Cu, Cr, Pb, Cd) in PM using a combination of colorimetric and electrochemical detection motifs. Detection limits of 0.1?g were achieved for colorimetric assays (appropriate for occupational exposures) while electrochemical assays provided detection limits of 0.1 ng (1 ppb solution levels; appropriate for ambient PM). We have also validated these techniques against standard reference methods with relatively good agreement. To complement these analytic techniques, our group has also developed a series of low-weight, low-burden personal air samplers. These air samplers feature micropumps that provide silent operation and a leap in energy efficiency that allows significant minimization. Initial prototypes weigh less than 150g, emit virtually no noise, and are about the size of a box of crayons. Production costs for these air sampling pumps are targeted for < $100, about ten times less than current competing technologies.
The global COVID-19 pandemic has created an urgent demand for accurate rapid point of care diagnostic tests. Antigen-based assays are suitably inexpensive and can be rapidly mass-produced, but sufficiently accurate performance requires highly-optimized antibodies and assay conditions. An automated liquid handling system, customized to handle lateral flow immunoassay (LFA) arrays, was used for high-throughput antibody screening of anti-nucleocapsid antibodies that will perform optimally on an LFA. Six hundred seventy-three anti-nucleocapsid antibody pairs were tested as both capture and detection reagents with the goal of finding those pairs that have the greatest affinity for unique epitopes of the nucleocapsid protein of SARS-CoV-2 while also performing optimally in an LFA format. In contrast to traditional antibody screening methods (e.g. ELISA, bio-layer interferometry), the methods described here integrate real-time LFA reaction kinetics and binding directly on nitrocellulose. We have identified several candidate antibody pairs that are suitable for further development of an LFA for SARS-CoV-2.
The release of metals and metal-containing compounds into the environment is a growing concern in developed and developing countries, as human exposure to metals is associated with adverse health effects in virtually every organ system. Unfortunately, quantifying metals in the environment is expensive; analysis costs using certified laboratories typically exceed $100/sample, making the routine analysis of toxic metals cost-prohibitive for applications such as occupational exposure or environmental protection. Here, we report on a simple, inexpensive technology with the potential to render toxic metals detection accessible for both the developing and developed world that combines colorimetric and electrochemical microfluidic paper-based analytical devices (mPAD) in a three-dimensional configuration. Unlike previous mPADs designed for measuring metals, the device reported here separates colorimetric detection on one layer from electrochemical detection on a different layer. Separate detection layers allows different chemistries to be applied to a single sample on the same device. To demonstrate the effectiveness of this approach, colorimetric detection is shown for Ni, Fe, Cu, and Cr and electrochemical detection for Pb and Cd. Detection limits as low as 0.12 μg (Cr) were achieved on the colorimetric layer while detection limits as low as 0.25 ng (Cd and Pb) were achieved on the electrochemical layer. Selectivity for the target analytes was demonstrated for common interferences. As an example of the device utility, particulate metals collected on air sampling filters were analyzed. Levels measured with the mPAD matched known values for the certified reference samples of collected particulate matter.
This paper reports a novel microfluidic device for characterizing low-copy enzymatic reactions via parallel mixing of picoliter-scale fluid volumes. Integrated polydimethylsiloxane (PDMS) microvalve arrays are used to control filling and mixing of a nanoliter-sized chamber with 1-9 picoliter-sized chambers. A proof-ofconcept enzymatic reaction was performed in which the device was loaded with the enzyme alkaline phosphatase (ALP) and the fluorogenic substrate fluorescein diphosphate (FDP). Fluorescence microscopy was used to observe the rate of fluorescence increase once enzyme and substrate solutions were allowed to diffusively mix.
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.
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.
This paper describes the synthesis of single-crystal nanowires of Pt directly on the surface of metal gauze made of Pt or W and demonstrates a more attractive approach for the growth of Pt nanowires on functional solid supports. There is evidence that the growth mechanism of the nanowires is correlated to the concentration of the Pt precursor and the surface roughness of the substrate. Electrochemical measurements indicate that the active surface area of the Pt nanowire-coated gauze is about 2−3 orders of magnitude greater than the pristine gauze. The results described herein demonstrate a new type of conductive support that can be used as active components for fuel cell applications and as an ideal 3D model catalyst.
