scholarly journals A modular paper-and-plastic device for tuberculosis nucleic acid amplification testing in limited-resource settings

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Navjot Kaur ◽  
Joy S. Michael ◽  
Bhushan J. Toley
Keyword(s):  
Nucleic Acid ◽  
Point Of Care ◽  
Limited Resource ◽  
High Specificity ◽  
Nucleic Acid Amplification ◽  
Analytical Sensitivity ◽  
Preparation Technique ◽  
Middle Income ◽  
Clinical Sensitivity ◽  
The Cost

Abstract We present a prototype for conducting rapid, inexpensive and point-of-care-compatible nucleic acid amplification tests (NAATs) for tuberculosis (TB). The fluorescent isothermal paper-and-plastic NAAT (FLIPP-NAAT) uses paper-based loop mediated isothermal amplification (LAMP) for DNA detection. The cost of materials required to build a 12-test-zone device is $0.88 and the cost of reagents per reaction is $0.43. An inexpensive imaging platform enables filter-free fluorescence detection of amplified DNA using a cell-phone camera. FLIPP-NAAT can be operated by an untrained user and only requires a regular laboratory incubator as ancillary equipment. All reagents can be dry-stored in the device, facilitating storage and transportation without cold chains. The device design is modular and the assay demonstrated high specificity to Mycobacterium tuberculosis (Mtb), analytical sensitivity of the order of 10 copies of Mtb gDNA, and tolerance to complex samples. The clinical sensitivity and specificity of sputum-based FLIPP NAAT tests were 100% (zero false negatives) and 68.75% (five false positives), respectively (N = 30), using Xpert MTB/RIF assay as the reference standard. FLIPP-NAAT has the potential to provide affordable and accessible molecular diagnostics for TB in low- and middle-income countries, when used in conjunction with an appropriate sample preparation technique. Although demonstrated for the detection of TB, FLIPP-NAAT is a platform technology for amplification of any nucleic acid sequence.

scholarly journals Limited-Resource Preparable Chitosan Magnetic Particles for Extracting Amplification-Ready Zeptomole-Range Nucleic Acid from Complex Biofluid

2021 ◽  
Author(s):  
Sayantan Tripathy ◽  
Arunansu Talukdar ◽  
Goutam Pramanik ◽  
P. V. Rajesh ◽  
Souradyuti Ghosh
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Magnetic Particles ◽  
Limited Resource ◽  
Nucleic Acid Amplification ◽  
Acid Extraction ◽  
Analytical Sensitivity ◽  
Nucleic Acid Extraction ◽  
Spin Column

<b>Layman Summary: </b>Nucleic acid extraction is a key prerequisite for any nucleic acid amplification test (NAAT) or isothermal NAAT (iNAAT) based molecular diagnosis assays.<b> </b>Existing methods utilizes spin column system for nucleic acid extraction which are unsuitable for limited resource settings. Our work explores two methods for chitosan coated magnetic particle preparation that can be executed within 6 h from commonly available chemicals with nothing but a magnetic stirrer and water bath and doable by a minimally trained person. We will also investigated the compatibility of the extracted nucleic acid with downstream NAATs such as real time LAMP, colorimetric LAMP, and real time PCR. In the process, we established the analytical sensitivity of the overall method.<div><br><div><b>Characterization methods</b>: SEM, XRD, EDX, FT-IR</div><div><br></div><div><b>Bioanalytical methods:</b> Real time LAMP, Colorimetric LAMP, Real time PCR</div></div>

scholarly journals Quantitative Isothermal Amplification on Paper Membranes using Amplification Nucleation Site Analysis

2022 ◽  
Author(s):  
Benjamin P. Sullivan ◽  
Yu-Shan Chou ◽  
Andrew T. Bender ◽  
Coleman D. Martin ◽  
Zoe G. Kaputa ◽  
...  
Keyword(s):  
Viral Load ◽  
Nucleic Acid ◽  
Infectious Diseases ◽  
Point Of Care ◽  
Nucleation Site ◽  
Isothermal Amplification ◽  
Nucleic Acid Amplification ◽  
Hiv Viral Load ◽  
Middle Income ◽  
Nucleation Sites

Quantitative nucleic acid amplification tests (qNAATs) are critical in treating infectious diseases, such as in HIV viral load monitoring or SARS-CoV-2 testing, in which viral load indicates viral suppression or infectivity. Quantitative PCR is the gold standard tool for qNAATs; however, there is a need to develop point-of-care (POC) qNAATs to manage infectious diseases in outpatient clinics, low- and middle-income countries, and the home. Isothermal amplification methods are an emerging tool for POC NAATs as an alternative to traditional PCR-based workflows. Previous works have focused on relating isothermal amplification bulk fluorescence signals to input copies of target nucleic acids for sample quantification with limited success. In this work, we show that recombinase polymerase amplification (RPA) reactions on paper membranes exhibit discrete fluorescent amplification nucleation sites. We demonstrate that the number of nucleation sites can be used to quantify HIV-1 DNA and RNA in less than 20 minutes. An image-analysis algorithm quantifies nucleation sites and determines the input nucleic acid copies in the range of 67-3,000 copies per reaction. We demonstrate a mobile phone-based system for image capture and onboard processing, illustrating that this method may be used at the point-of-care for qNAATs with minimal instrumentation.

scholarly journals Comparative performance of four nucleic acid amplification tests for SARS-CoV-2 virus

2020 ◽  
Author(s):  
Yujuan Xiong ◽  
Zhen-Zhen Li ◽  
Qi-Zhen Zhuang ◽  
Yan Chao ◽  
Fei Li ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Limit Of Detection ◽  
High Specificity ◽  
Nucleic Acid Amplification ◽  
Virus Concentration ◽  
Analytical Sensitivity ◽  
Comparative Performance ◽  
Consistency Test ◽  
Nucleic Acid Amplification Tests ◽  
Selection Of

AbstractCoronavirus disease 2019 (COVID-19) can be screened and diagnosed through the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by real-time reverse transcription polymerase chain reaction. SARS-CoV-2 nucleic acid amplification tests (NAATs) have been rapidly developed and quickly applied to clinical testing during the pandemic. However, studies evaluating the performance of these NAAT assays are limited. We evaluated the performance of four NAATs, which were marked by the Conformité Européenne and widely used in China during the pandemic. Results showed that the analytical sensitivity of the four assays was significantly lower than that claimed by the NAAT manufacturers. The limit of detection (LOD) of Daan, Sansure, and Hybribio NAATs was 3000 copies/mL, whereas the LOD of Bioperfectus NAATs was 4000 copies/mL. The results of the consistency test using 46 samples showed that Daan, Sansure, and Hybribio NAATs could detect the samples with a specificity of 100% (30/30) and a sensitivity of 100% (16 /16), whereas Bioperfectus NAAT detected the samples with a specificity of 100% (30/30) and a sensitivity 81.25% (13/16). The sensitivity of Bioperfectus NAAT was lower than that of the three other NAATs; this finding was consistent with the result that Bioperfectus NAAT had a higher LOD than the three other kinds of NAATs. The four above mentioned reagents presented high specificity; however, for the detection of the samples with low virus concentration, Bioperfectus reagent had the risk of missing detection. Therefore, the LOD should be considered in the selection of SARS-CoV-2 NAATs.

scholarly journals Limited-Resource Preparable Chitosan Magnetic Particles for Extracting Amplification-Ready Zeptomole-Range Nucleic Acid from Complex Biofluid

2021 ◽  
Author(s):  
Sayantan Tripathy ◽  
Arunansu Talukdar ◽  
Goutam Pramanik ◽  
P. V. Rajesh ◽  
Souradyuti Ghosh
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Magnetic Particles ◽  
Limited Resource ◽  
Nucleic Acid Amplification ◽  
Acid Extraction ◽  
Analytical Sensitivity ◽  
Nucleic Acid Extraction ◽  
Spin Column

<b>Layman Summary: </b>Nucleic acid extraction is a key prerequisite for any nucleic acid amplification test (NAAT) or isothermal NAAT (iNAAT) based molecular diagnosis assays.<b> </b>Existing methods utilizes spin column system for nucleic acid extraction which are unsuitable for limited resource settings. Our work explores two methods for chitosan coated magnetic particle preparation that can be executed within 6 h from commonly available chemicals with nothing but a magnetic stirrer and water bath and doable by a minimally trained person. We will also investigated the compatibility of the extracted nucleic acid with downstream NAATs such as real time LAMP, colorimetric LAMP, and real time PCR. In the process, we established the analytical sensitivity of the overall method.<div><br><div><b>Characterization methods</b>: SEM, XRD, EDX, FT-IR</div><div><br></div><div><b>Bioanalytical methods:</b> Real time LAMP, Colorimetric LAMP, Real time PCR</div></div>

scholarly journals The role of rapid diagnostic point of care IgG/IgM antibody tests in the diagnosis of SARS-CoV-2 infection

2020 ◽  
Vol 4 (1) ◽  
Author(s):  
Nishita Tripathi ◽  
Keyword(s):  
Nucleic Acid ◽  
Point Of Care ◽  
Clinical Importance ◽  
Antibody Test ◽  
High Specificity ◽  
Nucleic Acid Amplification ◽  
Serological Tests ◽  
Nucleic Acid Amplification Tests ◽  
Igm Antibody ◽  
Antibody Tests

Background: Current testing of symptomatic patients for SARS-CoV-2 involves the use of nucleic acid amplification tests, also known as genetic, RNA or PCR to detect viral RNA. The initial use of point-of-care (POC) antibody tests, also known as serological tests in the management of SARS-CoV-2 infection was limited. In this review, we determine the significance of POC antibody serological tests and explore their possible role in the diagnosis and management of patients infected with SARS-CoV-2 virus. Methods: A literature search was conducted in Google Scholar, PubMed, and Embase, and supplemented by searching the Center for Disease Control (CDC), and the Infectious Diseases Society of America (IDSA) websites. We identified 7 articles published in the last 6 months pertaining to the keywords. The sensitivity and specificity of the IgG/IgM antibody tests obtained from these studies were compared and used to determine the clinical importance of the rapid diagnostic antibody test in SARS-CoV-2 infection. Results: Through the literature review, it was found that POC diagnostic antibody tests can be used as an adjuvant with the nucleic acid amplification tests in determining both active and post-exposure antibodies. These rapid antibody IgG/IgM tests had high sensitivity, the ability of a test to correctly identify those with the disease, and high specificity, the ability of the test to correctly identify those without the disease. Conclusion: Emerging studies indicate the importance of POC antibody serological testing as an important diagnostic tool in the current SARS-CoV-2 pandemic. Considering the limitations of the molecular methods of testing, POC antibody tests can help reduce dependency on the molecular assays of testing when used in conjunction with them.

Limited-Resource Preparable Chitosan Magnetic Particles for Extracting Amplification-Ready Nucleic Acid from Complex Biofluid

The Analyst ◽  
2021 ◽  
Author(s):  
Sayantan Tripathy ◽  
Ashish Chalana ◽  
Arunansu Talukdar ◽  
P.V. Rajesh ◽  
Abhijit Saha ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Pathogen Detection ◽  
Magnetic Particles ◽  
Genetic Diseases ◽  
Limited Resource ◽  
Nucleic Acid Amplification Test ◽  
Nucleic Acid Amplification

Extraction and concentration of pure nucleic acid from complex biofluids is the prerequisite for nucleic acid amplification test (NAAT) applications in pathogen detection, biowarfare prevention, and genetic diseases. However, conventional...

Long-term dry storage of enzyme-based reagents for isothermal nucleic acid amplification in a porous matrix for use in point-of-care diagnostic devices

The Analyst ◽  
2020 ◽  
Vol 145 (21) ◽  
pp. 6875-6886 ◽  
Author(s):  
Sujatha Kumar ◽  
Ryan Gallagher ◽  
Josh Bishop ◽  
Enos Kline ◽  
Joshua Buser ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Glass Fiber ◽  
Point Of Care ◽  
Porous Matrix ◽  
Isothermal Amplification ◽  
Nucleic Acid Amplification ◽  
Isothermal Nucleic Acid Amplification ◽  
Dry Storage

Long-term dry storage of enzyme-based isothermal amplification reagents in glass fiber porous matrix for use in point-of-care devices.

scholarly journals 2158. Cost-Effectiveness and Budget Impact of a Point-of-Care Nucleic Acid Amplification Test for Diagnosis of Group A Streptococcal Pharyngitis in the United States

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S732-S732
Author(s):  
James Karichu ◽  
Mindy Cheng ◽  
Joanna Sickler ◽  
Julie Munakata ◽  
S Pinar Bilir ◽  
...  
Keyword(s):  
United States ◽  
Cost Effectiveness ◽  
Nucleic Acid ◽  
Budget Impact ◽  
Point Of Care ◽  
Cost Savings ◽  
The United States ◽  
Nucleic Acid Amplification ◽  
Sensitivity Analyses ◽  
Group A

Abstract Background Group A streptococcal (GAS) pharyngitis is common in the United States (US). Each year, approximately 12 million people seek medical care for pharyngitis, accounting for ~2% of ambulatory care visits. The gold standard method for diagnosing GAS is culture. However, because culture is time intensive, rapid antigen detection tests (RADTs), with or without culture confirmation, are commonly used. Although RADTs provide results quickly, test sensitivity has been shown to be sub-optimal, which can lead to inappropriate treatment decisions. Recently, highly sensitive point-of-care nucleic acid amplification tests (POC NAAT), such as the cobas® Liat® System, have emerged. The objective of this study was to evaluate the cost-effectiveness (CE) and budget impact (BI) of adopting POC NAAT compared with RADT+culture confirmation to diagnose GAS pharyngitis from the US third-party payer perspective. Methods A decision-tree economic model was developed in Microsoft Excel to quantify costs and clinical outcomes associated with POC NAAT and RADT+culture over a one-year period. All model inputs were derived from published literature and public databases. Model outputs included costs and clinical effects measured as quality-adjusted life days (QALDs) lost. One-way and probabilistic sensitivity analyses were performed to assess the impact of uncertainty on results. Results CE analysis showed that POC NAAT would cost $44 per patient compared with $78 with RADT+ culture. POC NAAT was associated with fewer QALDs lost relative to RADT+ culture. Therefore, POC NAAT may be considered the “dominant” strategy (i.e., lower costs and higher effectiveness). Findings were robust in sensitivity analyses. BI analysis showed that adopting POC NAAT for diagnosis of GAS could yield cost-savings of 0.3% vs. current budget over 3 years. This is due to savings associated with testing, GAS-related complications, antibiotic treatment and treatment-associated complication costs. Conclusion Results suggest that adopting POC NAAT to diagnose GAS would be considered cost-effective and yield cost-savings for US payers relative to RADT+culture. Access to POC NAAT would be important to optimize appropriate GAS diagnosis and treatment decisions. Disclosures All authors: No reported disclosures.

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