scholarly journals Heterogeneous Immunoassay Using Channels and Droplets in a Digital Microfluidic Platform

Micromachines ◽  
2019 ◽  
Vol 10 (2) ◽  
pp. 107 ◽  
Author(s):  
Yuguang Liu ◽  
Ian Papautsky
Keyword(s):  
Flow Rate ◽  
Retention Rate ◽  
Color Space ◽  
Magnetic Bead ◽  
Critical Parameters ◽  
Enzyme Linked Immunosorbent Assay ◽  
Discrete Sample ◽  
Wash Buffer ◽  
Heterogeneous Immunoassay ◽  
Digital Microfluidic

This work presents a heterogeneous immunoassay using the integrated functionalities of a channel and droplets in a digital microfluidic (DMF) platform. Droplet functionality in DMF allows for the programmable manipulation of discrete sample and reagent droplets in the range of nanoliters. Pressure-driven channels become advantageous over droplets when sample must be washed, as the supernatant can be thoroughly removed in a convenient and rapid manner while the sample is immobilized. Herein, we demonstrate a magnetic bead-based, enzyme-linked immunosorbent assay (ELISA) using ~60 nL of human interleukin-6 (IL-6) sample. The wash buffer was introduced in the form of a wall-less virtual electrowetting channel by a syringe pump at the flow rate of 10 μL/min with ~100% bead retention rate. Critical parameters such as sample wash flow rate and bead retention rate were optimized for reliable assay results. A colorimetric readout was analyzed in the International Commission on Illumination (CIE) color space without the need for costly equipment. The concepts presented in this work are potentially applicable in rapid neonatal disease screening using a finger prick blood sample in a DMF platform.

scholarly journals Technical viability of the YF MAC-HD ELISA kit for use in yellow fever-endemic regions

2021 ◽  
Vol 15 (6) ◽  
pp. e0009417
Author(s):  
Christin H. Goodman ◽  
Maurice Demanou ◽  
Mick Mulders ◽  
Jairo Mendez-Rico ◽  
Alison Jane Basile
Keyword(s):  
South America ◽  
Yellow Fever ◽  
Accurate Diagnosis ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Elisa Kit ◽  
Wash Buffer ◽  
Antibody Capture ◽  
Vaccination Campaigns ◽  
Arboviral Disease

Yellow fever (YF), an arboviral disease, affects an estimated 200,000 people and causes 30,000 deaths per year and recently has caused major epidemics in Africa and South America. Timely and accurate diagnosis of YF is critical for managing outbreaks and implementing vaccination campaigns. A YF immunoglobulin M (IgM) antibody-capture (MAC) enzyme-linked immunosorbent assay (ELISA) kit, the YF MAC-HD, was successfully introduced starting in 2018 to laboratories in Africa and South America. The YF MAC-HD kit can be performed in 3.5 hours, test up to 24 samples, and includes all reagents necessary to perform the test, except for water used to dilute wash buffer. In 2018 and 2019, a total of 56 laboratory personnel from 39 countries in Africa and South America were trained to use the kit during workshops, followed by take-home YF IgM proficiency testing (PT) exercises. Participants received either a 10- or 20-sample YF PT panel and performed testing using the YF MAC-HD kit. All countries obtained 90% or higher correct results. These results verified the technical viability and transferability of YF MAC-HD kit use for laboratories in YF-endemic countries.

scholarly journals Rab GTPase Mediating Regulation of NALP3 in Colorectal Cancer

Molecules ◽  
2020 ◽  
Vol 25 (20) ◽  
pp. 4834
Author(s):  
Gülçin Tezcan ◽  
Ekaterina E. Garanina ◽  
Margarita N. Zhuravleva ◽  
Shaimaa Hamza ◽  
Albert A. Rizvanov ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Vesicle Trafficking ◽  
Magnetic Bead ◽  
Dominant Negative ◽  
Enzyme Linked Immunosorbent Assay ◽  
Inflammasome Activation ◽  
Rab Gtpase ◽  
Rab Proteins ◽  
Caspase 1

The NALP3 inflammasome signaling contributes to inflammation within tumor tissues. This inflammation may be promoted by the vesicle trafficking of inflammasome components and cytokines. Rab5, Rab7 and Rab11 regulate vesicle trafficking. However, the role of these proteins in the regulation of inflammasomes remains largely unknown. To elucidate the role of these Rab proteins in inflammasome regulation, HCT-116, a colorectal cancer (CRC) cell line expressing pDsRed-Rab5 wild type (WT), pDsRed-Rab5 dominant-negative (DN), pDsRed-Rab7 WT, pDsRed-Rab7 DN, pDsRed-Rab11 WT and pDsRed-Rab11 DN were treated with lipopolysaccharide (LPS)/nigericin. Inflammasome activation was analyzed by measuring the mRNA expression of NLRP3, Pro-CASP1, RAB39A and Pro-IL-1β, conducting immunofluorescence imaging and western blotting of caspase-1 and analysing the secretion levels of IL-1β using enzyme-linked immunosorbent assay (ELISA). The effects of Rabs on cytokine release were evaluated using MILLIPLEX MAP Human Cytokine/Chemokine Magnetic Bead Panel-Premixed 41 Plex. The findings showed that LPS/nigericin-treated cells expressing Rab5-WT indicated increased NALP3 expression and secretion of the IL-1β as compared to Rab5-DN cells. Caspase-1 was localized in the nucleus and cytosol of Rab5-WT cells but was localized in the cytosol in Rab5-DN cells. There were no any effects of Rab7 and Rab11 expression on the regulation of inflammasomes. Our results suggest that Rab5 may be a potential target for the regulation of NALP3 in the treatment of the CRC inflammation.

scholarly journals Numerical Investigation of a Portable Incinerator: A Parametric Study

Processes ◽  
2020 ◽  
Vol 8 (8) ◽  
pp. 923
Author(s):  
Mohsen Saffari Pour ◽  
Ali Hakkaki-Fard ◽  
Bahar Firoozabadi
Keyword(s):  
Flow Rate ◽  
Fossil Fuels ◽  
Combustion Process ◽  
Critical Parameters ◽  
Mesh Quality ◽  
Hazardous Gases ◽  
Computational Fluid Dynamics Cfd ◽  
Reliable Design ◽  
Cooling Air ◽  
Numerical Approaches

The application of incinerators for the municipal solid waste (MSW) is growing due to the ability of such instruments to produce energy and, more specifically, reduce waste volume. In this paper, a numerical simulation of the combustion process with the help of the computational fluid dynamics (CFD) inside a portable (mobile) incinerator has been proposed. Such work is done to investigate the most critical parameters for a reliable design of a domestic portable incinerator, which is suitable for the Iranian food and waste culture. An old design of a simple incinerator has been used to apply the natural gas (NG), one of the available cheap fossil fuels in Iran. After that, the waste height, place of the primary burner, and the flow rate of the cooling air inside the incinerator, as the main parameters of the design, are investigated. A validation is also performed for the mesh quality test and the occurrence of the chemical reactions near the burner of the incinerator. Results proved that the numerical results have less than 5% error compared to the previous experimental and numerical approaches. In addition, results show that by moving the primary burner into the secondary chamber of the incinerator, the temperature and the heating ability of the incinerator could be affected dramatically. Moreover, it has been found that by increasing the flow rate of the cooling air inside the incinerator to some extent, the combustion process is improved and, on the other hand, by introducing more cooling air, the evacuation of the hazardous gases from the exhaust is also improved.

scholarly journals Critical parameters in the production of ceramic pot filters for household water treatment in developing countries

2014 ◽  
Vol 13 (2) ◽  
pp. 587-599 ◽  
Author(s):  
A. I. A. Soppe ◽  
S. G. J. Heijman ◽  
I. Gensburger ◽  
A. Shantz ◽  
D. van Halem ◽  
...  
Keyword(s):  
Developing Countries ◽  
Water Treatment ◽  
Removal Efficiency ◽  
Rice Husk ◽  
Flow Rate ◽  
Critical Parameters ◽  
Small Scale ◽  
Material Strength ◽  
Flow Rates ◽  
Pore Sizes

The need to improve the access to safe water is generally recognized for the benefit of public health in developing countries. This study's objective was to identify critical parameters which are essential for improving the performance of ceramic pot filters (CPFs) as a point-of-use water treatment system. Defining critical production parameters was also relevant to confirm that CPFs with high-flow rates may have the same disinfection capacity as pots with normal flow rates. A pilot unit was built in Cambodia to produce CPFs under controlled and constant conditions. Pots were manufactured from a mixture of clay, laterite and rice husk in a small-scale, gas-fired, temperature-controlled kiln and tested for flow rate, removal efficiency of bacteria and material strength. Flow rate can be increased by increasing pore sizes and by increasing porosity. Pore sizes were increased by using larger rice husk particles and porosity was increased with larger proportions of rice husk in the clay mixture. The main conclusions: larger pore size decreases the removal efficiency of bacteria; higher porosity does not affect the removal efficiency of bacteria, but does influence the strength of pots; flow rates of CPFs can be raised to 10–20 L/hour without a significant decrease in bacterial removal efficiency.

scholarly journals Development of Generic Immuno-Magnetic Bead-Based Enzyme-Linked Immunoassay for Ustiloxins in Rice Coupled with Enrichment

Toxins ◽  
2021 ◽  
Vol 13 (12) ◽  
pp. 907
Author(s):  
Yi Huang ◽  
Xiaoqian Tang ◽  
Lu Zheng ◽  
Junbin Huang ◽  
Qi Zhang ◽  
...  
Keyword(s):  
Magnetic Beads ◽  
High Efficiency ◽  
Detection Efficiency ◽  
Magnetic Bead ◽  
Hplc Method ◽  
Enzyme Linked Immunosorbent Assay ◽  
Common Structure ◽  
Site Monitoring ◽  
False Smut ◽  
Living Organisms

Ustiloxins are a group of mycotoxins produced by rice false smut pathogen. Previous studies have shown that the false smut balls contain six types of ustiloxins, and these toxins are toxic to living organisms. Thus, immunoassay for on-site monitoring of ustiloxins in rice is urgently required. The current immunoassays are only for detecting single ustiloxin, and they cannot meet the demand for synchronous and rapid detection of the group toxins. Therefore, this study designed and synthesized a generic antigen with ustiloxin G as material based on the common structure of the mycotoxins. Ustiloxin G was conjugated to two carrier proteins including bovine serum albumin (BSA) and ovalbvmin (OVA) by carbon diimide method. The mice were immunized with ustiloxin-G-BSA to generate the antibody serum, which was further purified to obtain the generic antibody against ustiloxins. The conjugated ustiloxin G-OVA and generic antibodies were used for establishing the enzyme-linked immunosorbent assay (ELISA) for ustiloxin detection and optimizing experiment conditions. The characterization of the antibody showed that the semi-inhibitory concentrations (IC50) of ustiloxin A, B, and G were 0.53, 0.34, and 0.06 µg/mL, respectively, and that their corresponding cross-reactivities were 11.9%, 18.4%, and 100%, respectively. To increase ELISA detection efficiency, generic antibody was combined with magnetic beads to obtain sensitive and class-specific immune-magnetic beads. Based on these immuno-magnetic beads, a high-efficiency enzyme-linked immunoassay method was developed for ustiloxin detection, whose sensitivity to ustiloxin A, B, and G was improved to 0.15 µg/mL, 0.14 µg/mL, and 0.04 µg/mL, respectively. The method accuracy was evaluated by spiking ustiloxin G as standard, and the spiked samples were tested by the immune-magnetic bead-based ELISA. The result showed the ustiloxin G recoveries ranged from 101.9% to 116.4% and were accepted by a standard HPLC method, indicating that our developed method would be promising for on-site monitoring of ustiloxins in rice.

Rapid Detection of Salmonella enteritidis in Pooled Liquid Egg Samples Using a Magnetic Bead-ELISA System

1995 ◽  
Vol 58 (9) ◽  
pp. 967-972 ◽  
Author(s):  
PETER S. HOLT ◽  
RICHARD K. GAST ◽  
CAM R. GREENE
Keyword(s):  
Salmonella Enteritidis ◽  
Magnetic Bead ◽  
Immunomagnetic Separation ◽  
Detection Methods ◽  
Enzyme Linked Immunosorbent Assay ◽  
Direct Plating ◽  
Elisa System ◽  
Low Concentrations ◽  
Standard Assay ◽  
Flagellar Proteins

An assay was developed to shorten the time necessary to detect Salmonella enteritidis (SE) in contaminated egg pools. The immunomagnetic separation (IMS)-based assay used the DynabeadsTM Anti-Salmonella, a magnetic bead with mouse anti-Salmonella antibodies affixed to the surface, to bind the SE in the egg pools. The bound SE were concentrated by a magnet and were detected via an enzyme-linked immunosorbent assay (ELISA) (IMS-ELISA) employing a monoclonal anti-SE flagellar proteins (flagellins) antibody. Following the ELISA, the beads were plated onto differential media (IMS-direct).The efficacy of the assay for detecting SE was compared with that of the standard assay, direct plating, in pooled egg samples spiked with low concentrations of SE and incubated at 37°C for 24 to 96 h. Conventional direct plating of egg samples required a total of 48 h before SE could be identified in egg pools, compared with 24 h for the IMS-ELISA. Plating of the beads (IMS-direct) to confirm the presence of SE required a further 24 h. The IMS-ELISA could detect SE at concentrations of 105 to 106 SE cells per ml, comparable to that shown previously for direct plating. The IMS-direct could detect SE at 104 SE cells per ml of egg pool. In egg pools initially contaminated with 10 SE cells per ml, the organism grew to levels by 24 h at 37°C where 100% of the pools were positive for SE by all three detection methods. In egg pools initially contaminated with 1 SE cell per ml, 61% of pools were detected by direct plating and IMS-ELISA and 72% were detected by IMS-direct. Similar detection frequencies were observed for a second SE isolate. The IMS-ELISA provides an SE detection rate comparable to direct plating but achieves the result 24 h sooner. The IMS-direct was the most sensitive means of detecting the SE.

On the selection of viscosity to suppress the Saffman–Taylor instability in a radially spreading annulus

2015 ◽  
Vol 782 ◽  
pp. 127-143 ◽  
Author(s):  
Tim H. Beeson-Jones ◽  
Andrew W. Woods
Keyword(s):  
Flow Rate ◽  
Viscosity Ratio ◽  
Early Time ◽  
Maximum Flow ◽  
Outer Radius ◽  
Relative Volume ◽  
Critical Parameters ◽  
Oil Well ◽  
Late Time ◽  
Azimuthal Mode

We examine the stability of a system with two radially spreading fronts in a Hele-Shaw cell in which the viscosity increases monotonically from the innermost to the outermost fluid. The critical parameters are identified as the viscosity ratio of the inner and outer fluids and the viscosity difference between the intermediate and outer fluids as a fraction of the viscosity difference between the inner and outer fluids. There is a minimum viscosity ratio of the inner and outer fluids above which, for each azimuthal mode, the system is stable to perturbations of that mode at any flow rate. This condition is directly analogous to the result for a single interface. Below this minimum ratio, the system may be stable at any flow rate early in the flow. However, once the inner radius reaches a critical fraction of the outer radius, this absolute stability ceases to apply owing to the coupling of the inner and outer interfaces. We determine the maximum flow rate, as a function of time, in order that all modes remain stable due to the effects of interfacial tension. These criteria for stability are then used to select the viscosity of the intermediate fluid so that a fixed volume of the intermediate and then inner fluid can be added to the system in the minimum time with the system remaining stable throughout. The optimal viscosity for this intermediate fluid depends on the relative volume of the inner and intermediate fluid and also on the overall viscosity ratio of the innermost fluid and the original fluid in the cell, with the balance being to suppress the early time instability of the outer interface and the late time instability of the inner interface. We discuss application of this approach to a problem of injection of treatment fluid in an oil well.

Method for Characterization of Passive Mechanical Filtration of Particles in Digital Microfluidic Devices

2014 ◽  
Author(s):  
Peter D. Dunning ◽  
Pierre E. Sullivan ◽  
Michael J. Schertzer
Keyword(s):  
Microfluidic Device ◽  
Vision System ◽  
Low Cost ◽  
Digital Microfluidics ◽  
Microfluidic Devices ◽  
Comb Filter ◽  
Enzyme Linked Immunosorbent Assay ◽  
Reaction Site ◽  
Digital Microfluidic

The ability to remove unbound biological material from a reaction site has applications in many biological protocols, such as those used to detect pathogens and biomarkers. One specific application where washing is critical is the Enzyme-Linked ImmunoSorbent Assay (ELISA). This protocol requires multiple washing steps to remove multiple reagents from a reaction site. Previous work has suggested that a passive mechanical comb filter can be used to wash particles in digital microfluidic devices. A method for the characterization of passive mechanical filtration of particles in Digital MicroFluidic (DMF) devices is presented in this work. In recent years there has been increased development of Lab-On-A-Chip (LOAC) devices for the automation and miniaturization of biological protocols. One platform for further research is in digital microfluidics. A digital microfluidic device can control the movement of pico-to nanoliter droplets of fluid using electrical signals without the use of pumps, valves, and channels. As such, fluidic pathways are not hardwired and the path of each droplet can be easily reconfigured. This is advantageous in biological protocols requiring the use of multiple reagents. Fabrication of these devices is relatively straight forward, since fluid manipulation is possible without the use of complex components. This work presents a method to characterize the performance of a digital microfluidic device using passive mechanical supernatant dilution via image analysis using a low cost vision system. The primary metric for performance of the device is particle retention after multiple passes through the filter. Repeatability of the process will be examined by characterizing performance of multiple devices using the same filter geometry. Qualitative data on repeatability and effectiveness of the dilution technique will also be attained by observing the ease with which the droplet disengages from the filter and by measuring the quantity of fluid trapped on the filter after each filtration step.

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