scholarly journals Massive Multiplexing Can Deliver a $1 Test for COVID-19

2020 ◽  
Author(s):  
Paul DN Hebert ◽  
Sean WJ Prosser ◽  
Natalia V Ivanova ◽  
Evgeny V Zakharov ◽  
Sujeevan Ratnasingham
Keyword(s):  
Large Scale ◽  
Low Cost ◽  
Rna Extraction ◽  
Respiratory Syndrome Virus ◽  
Major Advance ◽  
Rt Pcr ◽  
Screening Programs ◽  
Global Pandemic ◽  
Gene Coding ◽  
Large Scale Screening

ABSTRACTThe severe acute respiratory syndrome virus, SARS-CoV-2 (hereafter COVID-19), rapidly achieved global pandemic status, provoking large-scale screening programs in many nations. Their activation makes it imperative to identify methods that can deliver a diagnostic result at low cost. This paper describes an approach which employs sequence variation in the gene coding for its envelope protein as the basis for a scalable, inexpensive test for COVID-19. It achieves this by coupling a simple RNA extraction protocol with low-volume RT-PCR, followed by E-Gel screening and sequencing on high-throughput platforms to analyze 10,000 samples in a run. Slight modifications to the protocol could support screening programs for other known viruses and for viral discovery. Just as the $1,000 genome is transforming medicine, a $1 diagnostic test for viral and bacterial pathogens would represent a major advance for public health.

scholarly journals Rapid, Reliable and Robust approach for extraction-free RT-PCR based detection of SARS-CoV-2 in clinical setting to expedite large scale screening

2021 ◽  
Author(s):  
Abhilasha Dubey ◽  
Sanjay Upadhyay ◽  
Manjeet Mehta
Keyword(s):  
Large Scale ◽  
Target Genes ◽  
Low Cost ◽  
Diagnostic Methods ◽  
Analysis Method ◽  
Rt Pcr ◽  
Qpcr Method ◽  
Quantitative Results ◽  
Pooled Samples ◽  
Large Scale Screening

Rapid, reliable and robust method for the detection of SARS-CoV-2 is an indispensable need for diagnostics. The development of diagnostic methods will aid to address further waves of the pandemic potentially with rapid surveillance of disease and to allay the fears. To meet this challenge, we have developed a rapid RT-qPCR method for the detection of 3 target genes or confirmatory genes in less than 30 minutes. The assay showed 100% sensitivity and 100% specificity when tested on 120 samples. We compared a conventional extraction based method with extraction-free method, and then further reduced the run time of extraction free method. Additionally, we have validated our rapid RT-qPCR method for the assessment of pooled samples. We hereby propose a most reliable approach for the mass screening of samples with ease of operation at a low cost. Finally we designed a single tube analysis method which provides qualitative as well as quantitative results in minimum time.

scholarly journals Rapid, sensitive, full genome sequencing of Severe Acute Respiratory Syndrome Virus Coronavirus 2 (SARS-CoV-2)

2020 ◽  
Author(s):  
Clinton R Paden ◽  
Ying Tao ◽  
Krista Queen ◽  
Jing Zhang ◽  
Yan Li ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Sanger Sequencing ◽  
Respiratory Syndrome Virus ◽  
Full Genome ◽  
Miseq Sequencing ◽  
Genomic Information ◽  
Rt Pcr ◽  
Full Genome Sequencing ◽  
High Quality ◽  
Global Pandemic

AbstractSARS-CoV-2 recently emerged, resulting a global pandemic. Rapid genomic information is critical to understanding transmission and pathogenesis. Here, we describe validated protocols for generating high-quality full-length genomes from primary samples. The first employs multiplex RT-PCR followed by MinION or MiSeq sequencing. The second uses singleplex, nested RT-PCR and Sanger sequencing.

Impedance Screening for Preschool Children: State of the Art

1979 ◽  
Vol 88 (1) ◽  
pp. 56-65 ◽  
Author(s):  
Jack L. Paradise ◽  
Clyde G. Smith
Keyword(s):  
Preschool Children ◽  
Middle Ear ◽  
Large Scale ◽  
High Sensitivity ◽  
Cutoff Point ◽  
Screening Programs ◽  
Large Numbers ◽  
Middle Ear Disease ◽  
High Degree ◽  
Large Scale Screening

As a test for detecting middle ear disease among preschool children, tympanometry — as opposed to audiometry — has three advantageous attributes: a high degree of sensitivity, minimal need for subject cooperation, and total objectivity. For these reasons interest has arisen in tympanometry as a method for screening, i.e., identifying children with previously undetected middle ear disease. However, uncertainty persists concerning the importance of detecting apparently asymptomatic middle ear effusions, and concerning optimal methods, or even the advisability, of treating them. Further, the sensitivity and specificity of tympanometry depend on how the pass-fail cutoff point is defined. Defining this cutoff point so as to achieve high sensitivity may result in excessively low specificity, with the production of large numbers of false-positives who then become overreferrals. Data are presented to show how the validity of the test may be increased to some extent by attention to the gradient of “negative-pressure” tympanograms. At the present time, given the various aforementioned uncertainties, and with adequate validation as to the presence or absence of disease often lacking in reported studies of impedance screening in preschool populations, the cumulative results of these studies do not warrant embarking on large-scale screening programs. What is needed instead is additional research to explore the issue further.

scholarly journals Fast and low-cost detection of SARS-CoV-2 peptides by tandem mass spectrometry in clinical samples

2020 ◽  
Author(s):  
Karina Helena Morais Cardozo ◽  
Adriana Lebkuchen ◽  
Guilherme Goncalves Okai ◽  
Rodrigo Andrade Schuch ◽  
Luciana Godoy Viana ◽  
...  
Keyword(s):  
Real Time ◽  
Large Scale ◽  
Low Cost ◽  
Large Population ◽  
Virus Detection ◽  
Standard Test ◽  
Clinical Samples ◽  
Rt Pcr ◽  
Reverse Transcription Pcr ◽  
Fast Processing

Abstract The current outbreak of severe acute respiratory syndrome associated with coronavirus 2 (SARS-CoV-2) is pressing public health systems around the world, and large population testing is a key step to control this pandemic disease. Real-time reverse-transcription PCR (real-time RT-PCR) is the gold standard test for virus detection but the soaring demand for this test resulted in shortage of reagents and instruments, severely limiting its applicability to large-scale screening. To be used either as an alternative, or as a complement, to real-time RT-PCR testing, we developed a high-throughput targeted proteomics assay to detect SARS-CoV-2 proteins directly from clinical respiratory tract samples. Sample preparation was fully automated by using a modified magnetic particle-based proteomics approach implemented on a robotic liquid handler, enabling a fast processing of samples. The use of turbulent flow chromatography included four times multiplexed on-line sample cleanup and UPLC separation. MS/MS detection of three peptides from SARS-CoV-2 nucleoprotein and a 15N-labeled internal global standard was achieved within 2.5 min, enabling the analysis of more than 500 samples per day. The method was validated using 562 specimens previously analyzed by real-time RT-PCR and was able to detect over 83% of positive cases. No interference was found with samples from common respiratory viruses, including other coronaviruses (NL63, OC43, HKU1, and 229E). The strategy here presented has high sample stability and low cost and should be considered as an option to large population testing.

scholarly journals Large-scale implementation of pooled RNA extraction and RT-PCR for SARS-CoV-2 detection

2020 ◽  
Vol 26 (9) ◽  
pp. 1248-1253 ◽  
Author(s):  
R. Ben-Ami ◽  
A. Klochendler ◽  
M. Seidel ◽  
T. Sido ◽  
O. Gurel-Gurevich ◽  
...  
Keyword(s):  
Large Scale ◽  
Rna Extraction ◽  
Rt Pcr

scholarly journals Low-Cost Technology for the Prevention of Cervical Cancer in Cameroon

2020 ◽  
Vol 6 (Supplement_1) ◽  
pp. 8-8
Author(s):  
Nkegoum Blaise ◽  
Mboumtou Liliane
Keyword(s):  
Cervical Cancer ◽  
Predictive Value ◽  
Large Scale ◽  
Low Cost ◽  
Screening Method ◽  
Previous History ◽  
Pap Smears ◽  
Cervical Lesions ◽  
History Of ◽  
Large Scale Screening

PURPOSE Our aim was to assess the accuracy of visual inspection with acetic acid (VIA) as a screening method for cervical lesions. METHODS VIA and cytologic smears were carried out on the cervices of nonpregnant women age 30 to 60 years with no previous history of cervical cancer. Cervices with acetowhite lesions or positive Pap smears, as well as 1 in 10 negative cervices (control), were biopsied. RESULTS Of patients, 10,020 women were enrolled and 9,626 (96.1%) were screened. With screening, 9,534 patients (99.0%) had adequate cytology smears, 1,148 (11.9%) underwent colposcopy, and 3,486 biopsies were obtained, of which 1,056 were controls. Sensitivity of VIA was 70.4% versus 47.7%, specificity was 77.6% versus 94.2%, positive predictive value was 44.0% versus 67.2%, and negative predictive value was 91.3% versus 87.8% for Papanicolau test, respectively. CONCLUSION VIA has acceptable test qualities and is now well implemented as a large-scale screening method in Cameroon.

scholarly journals DNA-based carrier screening in primary healthcare: screening for aspartylglucosaminuria mutations in maternity health offices

1996 ◽  
Vol 42 (9) ◽  
pp. 1398-1404 ◽  
Author(s):  
M Hietala ◽  
P Aula ◽  
A C Syvänen ◽  
A Isoniemi ◽  
L Peltonen ◽  
...  
Keyword(s):  
Primary Healthcare ◽  
Large Scale ◽  
Solid Phase ◽  
Screening Program ◽  
Low Cost ◽  
Carrier Screening ◽  
Screening Programs ◽  
One Step ◽  
Laboratory Technology ◽  
Maternity Health

Abstract Large-scale genetic screening programs are complex enterprises in which ethical, technical, medical, and socioeconomic aspects have to be handled with professional expertise. Establishment of automated, relatively robust, and inexpensive laboratory techniques is one step of this path. Here a pilot carrier-screening program for the mutations causing aspartylglucosaminuria was carried out for pregnant women in primary care maternity health offices. Women (1975) were tested before their 12th week of pregnancy, and 31 heterozygotes were detected. The sampling was based on dried blood strips, facilitating convenient handling and inexpensive mailing to the laboratory. The mutation detection technique, solid-phase mini-sequencing simplified by the use of scintillation microplates and automated equipment, proved to be rapid, simple, inexpensive, and reliable, with a low repeat rate (2.5%). In conclusion, we found that good collaboration between the primary healthcare unit, the laboratory, and counseling experts, combined with modern laboratory technology, facilitate reliable low-cost genetic testing.

scholarly journals A simple and rapid method for isolating high-quality RNA from kenaf containing high polysaccharide and polyphenol contents

2020 ◽  
Author(s):  
Xiaofang Liao ◽  
Hongwei Li ◽  
Aziz Khan ◽  
Yanhong Zhao ◽  
Wenhuan Hou ◽  
...  
Keyword(s):  
Low Cost ◽  
Rna Extraction ◽  
Rna Isolation ◽  
Cost Effective ◽  
Spectrophotometric Analysis ◽  
Rt Pcr ◽  
High Quality ◽  
Time Saving ◽  
Ctab Method ◽  
Commercial Kits

AbstractThe isolation of high-quality RNA from kenaf is crucial for genetic and molecular biology studies. However, high levels of polysaccharide and polyphenol compounds in kenaf tissues could irreversibly bind to and coprecipitate with RNA, which complicates RNA extraction. In the present study, we proposed a simplified, time-saving and low-cost extraction method for isolating high quantities of high-quality RNA from several different kenaf tissues. RNA quality was measured for yield and purity, and the proposed protocol yielded high quantities of RNA (10.1-12.9 μg/g·FW). Spectrophotometric analysis showed that A260/280 ratios of RNA samples were in the range of 2.11 to 2.13, and A260/230 ratios were in the range of 2.04-2.24, indicating that the RNA samples were free of polyphenols, polysaccharides, and protein contaminants after isolation. The method of RNA extraction presented here was superior to the conventional CTAB method in terms of RNA isolation efficiency and was more sample-adaptable and cost-effective than commercial kits. Furthermore, to confirm downstream amenability, the high-quality RNA obtained from this method was successfully used for RT-PCR, real-time RT-PCR and Northern blot analysis. We provide an efficient and low-cost method for extracting high quantities of high-quality RNA from plants that are rich in polyphenols and polysaccharides, and this method was also validated for the isolation of high-quality RNA from other plants.

scholarly journals Fast and low-cost detection of SARS-CoV-2 peptides by tandem mass spectrometry in clinical samples

2020 ◽  
Author(s):  
Karina Helena Morais Cardozo ◽  
Adriana Lebkuchen ◽  
Guilherme Goncalves Okai ◽  
Rodrigo Andrade Schuch ◽  
Luciana Godoy Viana ◽  
...  
Keyword(s):  
Real Time ◽  
Large Scale ◽  
Low Cost ◽  
Large Population ◽  
Virus Detection ◽  
Standard Test ◽  
Clinical Samples ◽  
Rt Pcr ◽  
Reverse Transcription Pcr ◽  
Fast Processing

Abstract The current outbreak of severe acute respiratory syndrome associated with coronavirus 2 (SARS-CoV-2) is pressing public health systems around the world, and large population testing is a key step to control this pandemic disease. Real-time reverse-transcription PCR (real-time RT-PCR) is the gold standard test for virus detection but the soaring demand for this test resulted in shortage of reagents and instruments, severely limiting its applicability to large-scale screening. To be used either as an alternative, or as a complement, to real-time RT-PCR testing, we developed a high-throughput targeted proteomics assay to detect SARS-CoV-2 proteins directly from clinical respiratory tract samples. Sample preparation was fully automated by using a modified magnetic particle-based proteomics approach implemented on a robotic liquid handler, enabling a fast processing of samples. The use of turbulent flow chromatography included four times multiplexed on-line sample cleanup and UPLC separation. MS/MS detection of three peptides from SARS-CoV-2 nucleoprotein and a 15N-labeled internal global standard was achieved within 2.5 min, enabling the analysis of more than 500 samples per day. The method was validated using 562 specimens previously analyzed by real-time RT-PCR and was able to detect over 83% of positive cases. No interference was found with samples from common respiratory viruses, including other coronaviruses (NL63, OC43, HKU1, and 229E). The strategy here presented has high sample stability and low cost and should be considered as an option to large population testing.

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