scholarly journals Oil immersed lossless total analysis system for integrated RNA extraction and detection of SARS-CoV-2

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Duane S. Juang ◽  
Terry D. Juang ◽  
Dawn M. Dudley ◽  
Christina M. Newman ◽  
Molly A. Accola ◽  
...  
Keyword(s):  
Infectious Disease ◽  
Large Scale ◽  
Rna Extraction ◽  
Clinical Samples ◽  
Nasopharyngeal Swab ◽  
Acid Extraction ◽  
Total Analysis System ◽  
Total Analysis ◽  
Analysis System ◽  
Disease Testing

AbstractThe COVID-19 pandemic exposed difficulties in scaling current quantitative PCR (qPCR)-based diagnostic methodologies for large-scale infectious disease testing. Bottlenecks include lengthy multi-step processes for nucleic acid extraction followed by qPCR readouts, which require costly instrumentation and infrastructure, as well as reagent and plastic consumable shortages stemming from supply chain constraints. Here we report an Oil Immersed Lossless Total Analysis System (OIL-TAS), which integrates RNA extraction and detection onto a single device that is simple, rapid, cost effective, and requires minimal supplies and infrastructure to perform. We validated the performance of OIL-TAS using contrived SARS-CoV-2 viral particle samples and clinical nasopharyngeal swab samples. OIL-TAS showed a 93% positive predictive agreement (n = 57) and 100% negative predictive agreement (n = 10) with clinical SARS-CoV-2 qPCR assays in testing clinical samples, highlighting its potential to be a faster, cheaper, and easier-to-deploy alternative for infectious disease testing.

scholarly journals Oil Immersed Lossless Total Analysis System (OIL-TAS): Integrated RNA Extraction and Detection for SARS-CoV-2 Testing

2020 ◽  
Author(s):  
Duane S. Juang ◽  
Terry D. Juang ◽  
Dawn M. Dudley ◽  
Christina M. Newman ◽  
Thomas C. Friedrich ◽  
...  
Keyword(s):  
Infectious Disease ◽  
Large Scale ◽  
Rna Extraction ◽  
Cost Effective ◽  
Acid Extraction ◽  
Input Concentration ◽  
Total Analysis System ◽  
Total Analysis ◽  
Analysis System ◽  
Disease Testing

AbstractThe coronavirus disease 2019 (COVID-19) pandemic exposed difficulties in scaling current quantitative PCR (qPCR)-based diagnostic methodologies for large-scale infectious disease testing. Bottlenecks include the lengthy multi-step process of nucleic acid extraction followed by qPCR readouts, which require costly instrumentation and infrastructure, as well as reagent and plastic consumable shortages stemming from supply chain constraints. Here we report a novel Oil Immersed Lossless Total Analysis System (OIL-TAS), which integrates RNA extraction and detection onto a single device that is simple, rapid, cost effective, uses minimal supplies and requires reduced infrastructure to perform. We validated the performance of OIL-TAS using contrived samples containing inactivated SARS-CoV-2 viral particles, which show that the assay can reliably detect an input concentration of 10 copies/μL and sporadically detect down to 1 copy/μL. The OIL-TAS method can serve as a faster, cheaper, and easier-to-deploy alternative to current qPCR-based methods for infectious disease testing.

scholarly journals Comparison Between a Standard and SalivaDirect RNA Extraction Protocol for Molecular Diagnosis of SARS-CoV-2 Using Nasopharyngeal Swab and Saliva Clinical Samples

2021 ◽  
Vol 9 ◽  
Author(s):  
Sofía N. Rodríguez Flores ◽  
Luis Mario Rodríguez-Martínez ◽  
Bernardita L. Reyes-Berrones ◽  
Nadia A. Fernández-Santos ◽  
Elthon J. Sierra-Moncada ◽  
...  
Keyword(s):  
Rna Extraction ◽  
Proteinase K ◽  
Clinical Samples ◽  
Nasopharyngeal Swab ◽  
Acid Extraction ◽  
Sample Collection ◽  
Extraction Protocol ◽  
Assay Performance ◽  
Two Samples ◽  
Oropharyngeal Swab

During the COVID-19 pandemic, a certified laboratory of Tamaulipas, Mexico has processed over 100,000 samples of COVID-19 suspected patients, working a minimum of 100 tests daily. Thus, it would be beneficial for such certified laboratories nationwide to reduce the time and cost involved in performing the diagnosis of COVID-19, from sample collection, transportation to local lab, processing of samples, and data acquisition. Here, 30 nasopharyngeal swab and saliva samples from the same COVID-19 individuals were assessed by a standard nucleic acid extraction protocol, including protein lysis with proteinase K followed by binding to column, washing, and elution, and by the SalivaDirect protocol based on protein lysis, skipping the other steps to reduce processing time and costs. The genomic RNA was amplified using a SARS-CoV-2 Real-Time PCR kit. A variation (P > 0.05) in the 95% CIs = 72.6%–96.7% was noted by using the SalivaDirect protocol and saliva samples (sensitivity of 88.2%) in comparison to those of standard protocol with oropharyngeal swab samples (95% CIs = 97.5%–100%; sensitivity of 100%) as reported elsewhere. However, when using nasopharyngeal swab samples in the SalivaDirect protocol (sensitivity of 93.6%; 95% CIs = 79.2%–99.2%), it was in concordance (P < 0.05) with those of the standard one. The logical explanation to this was that two samples with Ct values of 38, and 40 cycles for gene E produced two false negatives in the SalivaDirect protocol in relation to the standard one; thus, there was a reduction of the sensitivity of 6.4% in the overall assay performance.

scholarly journals Evaluation of an Automated High-Throughput Liquid-Based RNA Extraction Platform on Pooled Nasopharyngeal or Saliva Specimens for SARS-CoV-2 RT-PCR

Viruses ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 615
Author(s):  
Allen Wing-Ho Chu ◽  
Cyril Chik-Yan Yip ◽  
Wan-Mui Chan ◽  
Anthony Chin-Ki Ng ◽  
Dream Lok-Sze Chan ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
High Throughput ◽  
Rna Extraction ◽  
Nasopharyngeal Swab ◽  
Acid Extraction ◽  
Rt Pcr ◽  
Nucleic Acid Extraction ◽  
Suitable Alternative ◽  
Pcr Inhibitors ◽  
Pcr Targeting

SARS-CoV-2 RT-PCR with pooled specimens has been implemented during the COVID-19 pandemic as a cost- and manpower-saving strategy for large-scale testing. However, there is a paucity of data on the efficiency of different nucleic acid extraction platforms on pooled specimens. This study compared a novel automated high-throughput liquid-based RNA extraction (LRE) platform (PHASIFYTM) with a widely used magnetic bead-based total nucleic acid extraction (MBTE) platform (NucliSENS® easyMAG®). A total of 60 pools of nasopharyngeal swab and 60 pools of posterior oropharyngeal saliva specimens, each consisting of 1 SARS-CoV-2 positive and 9 SARS-CoV-2 negative specimens, were included for the comparison. Real-time RT-PCR targeting the SARS-CoV-2 RdRp/Hel gene was performed, and GAPDH RT-PCR was used to detect RT-PCR inhibitors. No significant differences were observed in the Ct values and overall RT-PCR positive rates between LRE and MBTE platforms (92.5% (111/120] vs 90% (108/120]), but there was a slightly higher positive rate for LRE (88.3% (53/60]) than MBTE (81.7% (49/60]) among pooled saliva. The automated LRE method is comparable to a standard MBTE method for the detection of SAR-CoV-2 in pooled specimens, providing a suitable alternative automated extraction platform. Furthermore, LRE may be better suited for pooled saliva specimens due to more efficient removal of RT-PCR inhibitors.

scholarly journals Direct diagnostic testing of SARS-CoV-2 without the need for prior RNA extraction

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Shan Wei ◽  
Esther Kohl ◽  
Alexandre Djandji ◽  
Stephanie Morgan ◽  
Susan Whittier ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Point Of Care ◽  
Diagnostic Testing ◽  
Rna Extraction ◽  
Cost Effective ◽  
Clinical Samples ◽  
Nasopharyngeal Swab ◽  
Percentage Agreement ◽  
Testing Method ◽  
Viral Transport

AbstractThe COVID-19 pandemic has resulted in an urgent need for a rapid, point of care diagnostic testing that could be rapidly scaled on a worldwide level. We developed and tested a highly sensitive and robust assay based on reverse transcription loop mediated isothermal amplification (RT-LAMP) that uses readily available reagents and a simple heat block using contrived spike-in and actual clinical samples. RT-LAMP testing on RNA-spiked samples showed a limit of detection (LoD) of 2.5 copies/μl of viral transport media. RT-LAMP testing directly on clinical nasopharyngeal swab samples in viral transport media had an 85% positive percentage agreement (PPA) (17/20), and 100% negative percentage agreement (NPV) and delivered results in 30 min. Our optimized RT-LAMP based testing method is a scalable system that is sufficiently sensitive and robust to test for SARS-CoV-2 directly on clinical nasopharyngeal swab samples in viral transport media in 30 min at the point of care without the need for specialized or proprietary equipment or reagents. This cost-effective and efficient one-step testing method can be readily available for COVID-19 testing world-wide, especially in resource poor settings.

Detection of Directivity of Fluorescence From a Mixed Specimen Which Consists of Micro Particles Attached Different Fluorescent Substances Using a Light Waveguide Incorporated Optical TAS

2018 ◽  
Author(s):  
Toshifumi Ohkubo ◽  
Nobuyuki Terada ◽  
Yoshikazu Yoshida
Keyword(s):  
Microfluidic Channel ◽  
Laser Source ◽  
Microfluidic Channels ◽  
Fluorescent Substance ◽  
Total Analysis System ◽  
Micro Particles ◽  
Total Analysis ◽  
Light Waveguide ◽  
Analysis System ◽  
Near Future

A resin-based optical total analysis system (O-TAS) which consists both of microfluidic channels and light waveguides [1] is thought to be one of the most promising components in developing a “ubiquitous human healthcare system” in the near future. Along with this technology trend, we have already developed a transparent epoxy-resin-based optical TAS chip which has a specially prepared light waveguide structure of radially arranged configuration at an intersection portion with a microfluidic channel, in order to detect directivity of fluorescence from fluorescent substance attached micro particles [2],[3]. Schematic diagram of the optical TAS is shown in Figure 1. In the latest research, utilizing an AC modulated laser source and time-series averaging function on detected signal waveforms, we could have successfully obtained directivities of fluorescence from 5-μm-diameter particles with higher signal to noise (S/N) ratio [3].

Micro Particle Imaging Velocimetry Measurements in High Aspect Ratio Passive Microfluidic Components

2005 ◽  
Author(s):  
Paul Chiarot ◽  
Pierre Sullivan ◽  
Ridha Ben Mrad
Keyword(s):  
Aspect Ratio ◽  
High Aspect Ratio ◽  
Particle Imaging Velocimetry ◽  
Passive Component ◽  
Total Analysis System ◽  
Micro Piv ◽  
Micro Total Analysis Systems ◽  
Total Analysis ◽  
Analysis System ◽  
Particle Imaging

In this work, micro particle imaging velocimetry (micro-PIV) was performed on the fundamental components of a micro total analysis system. Specifically, high aspect ratio passive valves and mixers were designed, fabricated, and characterized. The components were built using Micralyne Protolyne technology on a glass substrate and operated at reasonably achievable pressures. The flows through the components were analyzed both qualitatively and quantitatively with the goal of developing a more complete understanding of internal device performance. Using the results of the micro-PIV developed velocity fields it was found that the high aspect ratio passive valves are able to perform at reasonably achievable pressures. However, it was determined that the high aspect ratio passive mixers offer limited performance enhancements because of the low Reynolds number flows. The results of this work contribute to the understanding of passive component operation and address some of the challenges associated with developing completely integrated micro total analysis systems that use passive devices.

scholarly journals Miniaturization of fluorescence sensing in optofluidic devices

2020 ◽  
Vol 24 (9) ◽  
Author(s):  
Daniel Măriuţa ◽  
Stéphane Colin ◽  
Christine Barrot-Lattes ◽  
Stéphane Le Calvé ◽  
Jan G. Korvink ◽  
...  
Keyword(s):  
Lab On A Chip ◽  
Time Data ◽  
Fluorescence Sensing ◽  
Successful Development ◽  
Total Analysis System ◽  
Total Analysis ◽  
Integration Strategies ◽  
Analysis System ◽  
On Chip ◽  
Industrial Sensing

Abstract Successful development of a micro-total-analysis system (µTAS, lab-on-a-chip) is strictly related to the degree of miniaturization, integration, autonomy, sensitivity, selectivity, and repeatability of its detector. Fluorescence sensing is an optical detection method used for a large variety of biological and chemical assays, and its full integration within lab-on-a-chip devices remains a challenge. Important achievements were reported during the last few years, including improvements of previously reported methodologies, as well as new integration strategies. However, a universal paradigm remains elusive. This review considers achievements in the field of fluorescence sensing miniaturization, starting from off-chip approaches, representing miniaturized versions of their lab counter-parts, continuing gradually with strategies that aim to fully integrate fluorescence detection on-chip, and reporting the results around integration strategies based on optical-fiber-based designs, optical layer integrated designs, CMOS-based fluorescence sensing, and organic electronics. Further successful development in this field would enable the implementation of sensing networks in specific environments that, when coupled to Internet-of-Things (IoT) and artificial intelligence (AI), could provide real-time data collection and, therefore, revolutionize fields like health, environmental, and industrial sensing.

Water glass bonding for micro-total analysis system

2002 ◽  
Vol 81 (2-3) ◽  
pp. 187-195 ◽  
Author(s):  
Takeshi Ito ◽  
Kazuharu Sobue ◽  
Seishiro Ohya
Keyword(s):  
Water Glass ◽  
Micro Total Analysis System ◽  
Total Analysis System ◽  
Total Analysis ◽  
Analysis System
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