scholarly journals Fast Detection of SARS-CoV-2 RNA Directly from Respiratory Samples Using a Loop-Mediated Isothermal Amplification (LAMP) Test

Viruses ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 801
Author(s):  
Olympia E. Anastasiou ◽  
Caroline Holtkamp ◽  
Miriam Schäfer ◽  
Frieda Schön ◽  
Anna Maria Eis-Hübinger ◽  
...  
Keyword(s):  
Predictive Value ◽  
Lamp Assay ◽  
Detection Methods ◽  
Analytical Sensitivity ◽  
Rt Pcr ◽  
Viral Transport Medium ◽  
Rapid Tests ◽  
Sous Vide ◽  
Antigen Tests ◽  
Isothermal Assay

The availability of simple SARS-CoV-2 detection methods is crucial to contain the COVID-19 pandemic. This study examined whether a commercial LAMP assay can reliably detect SARS-CoV-2 genomes directly in respiratory samples without having to extract nucleic acids (NA) beforehand. Nasopharyngeal swabs (NPS, n = 220) were tested by real-time reverse transcription (RT)-PCR and with the LAMP assay. For RT-PCR, NA were investigated. For LAMP, NA from 26 NPS in viral transport medium (VTM) were tested. The other 194 NPS were analyzed directly without prior NA extraction (140 samples in VTM; 54 dry swab samples stirred in phosphate buffered saline). Ten NPS were tested directly by LAMP using a sous-vide cooking unit. The isothermal assay demonstrated excellent specificity (100%) but moderate sensitivity (68.8%), with a positive predictive value of 1 and a negative predictive value of 0.65 for direct testing of NPS in VTM. The use of dry swabs, even without NA extraction, improved the analytical sensitivity; up to 6% of samples showed signs of inhibition. LAMP could be performed successfully with a sous-vide cooking unit. This technique is very fast, requires little laboratory resources, and can replace rapid antigen tests or verify reactive rapid tests on-site.

scholarly journals Analytical Sensitivity of the Abbott BinaxNOW COVID-19 Ag Card

2020 ◽  
Vol 59 (3) ◽  
Author(s):  
Garrett A. Perchetti ◽  
Meei-Li Huang ◽  
Margaret G. Mills ◽  
Keith R. Jerome ◽  
Alexander L. Greninger
Keyword(s):  
Reverse Transcription ◽  
Limit Of Detection ◽  
Digital Pcr ◽  
Detection Methods ◽  
Clinical Samples ◽  
Analytical Sensitivity ◽  
Qrt Pcr ◽  
Viral Transport Medium ◽  
Antigen Tests ◽  
Antigen Testing

ABSTRACT Multiple rapid antigen (Ag) tests for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have recently received emergency-use authorization (EUA) from the U.S. Food and Drug Administration (FDA). Although less sensitive than molecular detection methods, rapid antigen testing offers the potential for inexpensive, quick, decentralized testing. Robust analytical sensitivity data in comparison to reverse transcription-quantitative PCR (qRT-PCR) are currently lacking for many rapid antigen tests. Here, we evaluated the analytical sensitivity of the Abbott BinaxNOW COVID-19 Ag card using SARS-CoV-2-positive clinical specimens quantified by reverse transcription-droplet digital PCR (RT-ddPCR) and multiple FDA EUA qRT-PCR platforms using RNA standards. Initial and confirmatory limits of detection for the BinaxNOW COVID-19 Ag card were determined to be equivalent to 4.04 × 104 to 8.06 × 104 copies/swab. We further confirmed this limit of detection with 72 additional clinical samples positive for SARS-CoV-2 in either phosphate-buffered saline or viral transport medium. One hundred percent of samples with viral loads of >40,000 copies/swab were detected by rapid antigen testing. These data indicate that the BinaxNOW COVID-19 Ag card has an analytical sensitivity approximately equivalent to a generic qRT-PCR cycle threshold (CT) value of 29 to 30.

A prospective nationwide observational study of agreement of antigen tests on oral pharyngeal swabs or less invasive testing with RT-qPCR, for detecting SARS-CoV-2 in adults: Protocol description (Preprint)

2021 ◽  
Author(s):  
Uffe Vest Schneider ◽  
Jenny Dahl Knudsen ◽  
Anders Koch ◽  
Nikolai Søren Kirkby ◽  
Jan Gorm Lisby
Keyword(s):  
Confidence Intervals ◽  
Predictive Value ◽  
Large Scale ◽  
Reference Method ◽  
Lateral Flow ◽  
Clinical Samples ◽  
Sampling Location ◽  
Analytical Sensitivity ◽  
Clinical Sensitivity ◽  
Antigen Tests

BACKGROUND The SARS-CoV-2 pandemic has resulted in an unprecedented level of world-wide testing for epidemiologic and diagnostic purposes, and due to the extreme need for tests, the gold standard reverse transcription polymerase chain reaction (RT-qPCR) testing capacity has been unable to meet the overall global testing demand. Consequently, although current literature has shown the sensitivity of rapid antigen tests (RATs) to be inferior to RT-qPCR, RATs have been implemented on a large scale without solid data on performance. OBJECTIVE This study will compare analytical and clinical sensitivities and specificities of 50 lateral flow or laboratory based RATs and three Strand Invasion Based Amplification (SIBA)-rt-PCR tests from 30 manufacturers to RT-qPCR on samples obtained from the deep oropharynx. In addition, the study will compare sensitivities and specificities of the included RATs as well as RT-qPCR on clinical samples obtained from the deep oropharynx, anterior nasal cavity, saliva, deep nasopharynx and expired air to RT-qPCR from deep oropharyngeal samples. METHODS In the prospective part of the study, 200 individuals found SARS-CoV-2 positive and 200 individuals found SARS-CoV-2 negative by routine RT-qPCR testing will be re-tested with each RAT applying RT-qPCR as the reference method. In the retrospective part of the study, 304 deep oropharyngeal cavity swabs divided into four groups based on RT-qPCR Cq levels will be tested by each RAT. RESULTS The results will be reported in several manuscripts with different aims. The first manuscript will report retrospective (analytical sensitivity, overall and stratified into different Cq range groups) and prospective (clinical sensitivity) data for RATs with RT-qPCR results as the reference method. The second manuscript will report results for RAT based on anatomical sampling location. The third manuscript will compare different anatomical sampling locations by RT-qPCR testing. The fourth manuscript will focus on RATs that rely on central laboratory testing. Test from four different manufactures will be compared for analytical performance data on retrospective deep oropharyngeal swab samples. The fifth manuscript will report the results of four RATs applied both as professional use and as self-test. The last manuscript will report the results from two breath tests participating in the study. Comparison of sensitivity and specificity between RATs will be done using McNemar for paired samples and chi-squared test for unpaired samples. Comparison of PPV and NPV between RATs will be done by bootstrap test. 95 % confidence intervals for sensitivity, specificity, positive predictive value and negative predictive value are calculated as bootstrap confidence intervals CONCLUSIONS The study will compare the sensitivities of a large number of RATs for SARS-CoV-2 compared to RT-qPCR and will address whether lateral flow based RATs test differ significantly from laboratory based RATS. The anatomical test location for both RAT and RT-qPCR will be compared. CLINICALTRIAL ClinicalTrials.gov NCT04913116

scholarly journals Rapid isothermal amplification and portable detection system for SARS-CoV-2

2020 ◽  
Vol 117 (37) ◽  
pp. 22727-22735 ◽  
Author(s):  
Anurup Ganguli ◽  
Ariana Mostafa ◽  
Jacob Berger ◽  
Mehmet Y. Aydin ◽  
Fu Sun ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Detection System ◽  
Alternative Pathway ◽  
Rna Extraction ◽  
Lamp Assay ◽  
Isothermal Amplification ◽  
Clinical Samples ◽  
Complete Agreement ◽  
Rt Pcr ◽  
Viral Transport Medium

The COVID-19 pandemic provides an urgent example where a gap exists between availability of state-of-the-art diagnostics and current needs. As assay protocols and primer sequences become widely known, many laboratories perform diagnostic tests using methods such as RT-PCR or reverse transcription loop mediated isothermal amplification (RT-LAMP). Here, we report an RT-LAMP isothermal assay for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus and demonstrate the assay on clinical samples using a simple and accessible point-of-care (POC) instrument. We characterized the assay by dipping swabs into synthetic nasal fluid spiked with the virus, moving the swab to viral transport medium (VTM), and sampling a volume of the VTM to perform the RT-LAMP assay without an RNA extraction kit. The assay has a limit of detection (LOD) of 50 RNA copies per μL in the VTM solution within 30 min. We further demonstrate our assay by detecting SARS-CoV-2 viruses from 20 clinical samples. Finally, we demonstrate a portable and real-time POC device to detect SARS-CoV-2 from VTM samples using an additively manufactured three-dimensional cartridge and a smartphone-based reader. The POC system was tested using 10 clinical samples, and was able to detect SARS-CoV-2 from these clinical samples by distinguishing positive samples from negative samples after 30 min. The POC tests are in complete agreement with RT-PCR controls. This work demonstrates an alternative pathway for SARS-CoV-2 diagnostics that does not require conventional laboratory infrastructure, in settings where diagnosis is required at the point of sample collection.

scholarly journals Sensitivity of rapid antigen testing and RT-PCR performed on nasopharyngeal swabs versus saliva samples in COVID-19 hospitalized patients: results of a prospective comparative trial (RESTART)

2021 ◽  
Author(s):  
Antonios Kritikos ◽  
Giorgia Caruana ◽  
René Brouillet ◽  
John-Paul Miroz ◽  
Abed-Maillard Samia ◽  
...  
Keyword(s):  
Comparative Trial ◽  
Hospitalized Patients ◽  
Rt Pcr ◽  
Duration Of Symptoms ◽  
Non Invasive ◽  
Viral Transport Medium ◽  
One Step ◽  
Antigen Tests ◽  
Low Sensitivity ◽  
Antigen Testing

AbstractObjectivesSaliva sampling could serve as an alternative non-invasive sample for SARS-CoV-2 diagnosis while rapid antigen testing (RAT) might help to mitigate the shortage of reagents sporadically encountered with RT-PCR. Thus, in the RESTART study we compared antigen and RT-PCR testing methods on nasopharyngeal (NP) swabs and salivary samples.MethodsWe conducted a prospective observational study among COVID-19 hospitalized patients between 10th December 2020 and 1st February 2021. Paired saliva and NP samples were investigated by RT-PCR (Cobas 6800, Roche-Switzerland) and by two rapid antigen tests: One Step Immunoassay Exdia® COVID-19 Ag (Precision Biosensor, Korea) and Standard Q® COVID-19 Rapid Antigen Test (Roche-Switzerland).ResultsA total of 58 paired NP-saliva specimens were collected. Thirty-two of 58 (55%) patients were hospitalized in the intensive care unit and the median duration of symptoms was 11 days (IQR 5-19). NP and salivary RT-PCR exhibited sensitivity of 98% and 69% respectively whereas the specificity of these RT-PCRs assays were of 100%. NP RAT exhibited much lower diagnostic performances with sensitivities of 35% and 41% for the Standard Q® and Exdia® assays respectively, when a wet-swab approach was used (i.e. when the swab was diluted in the viral transport medium (VTM) before testing). The sensitivity of the dry-swab approach was slightly better (47%). These antigen tests exhibited very low sensitivity (4 and 8%) when applied to salivary swabs.ConclusionsNasopharyngeal RT-PCR is the most accurate test for COVID-19 diagnosis in hospitalized patients. RT-PCR on salivary samples may be used when nasopharyngeal swabs are contraindicated. RAT are not appropriate for hospitalized patients.

Performance evaluation of an automated SARS-CoV-2 Ag test for the diagnosis of COVID-19 infection on nasopharyngeal swabs

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Cédric Hartard ◽  
Sibel Berger ◽  
Thomas Josse ◽  
Evelyne Schvoerer ◽  
Hélène Jeulin
Keyword(s):  
Performance Evaluation ◽  
Negative Predictive Value ◽  
Symptom Onset ◽  
Predictive Value ◽  
Rt Pcr ◽  
Infected People ◽  
Antigen Tests

Abstract Objectives The detection of SARS-CoV-2 in infected people is a key tool to help in controlling COVID-19 pandemic. Like rapid antigenic tests, automated antigen tests, that present the advantage of a higher throughput flow, may be of interest. The LIAISON® SARS-CoV-2 Ag test was evaluated for the quantification of SARS-CoV-2 nucleocapsid antigen in nasopharyngeal swabs by comparison to RT-PCR. Methods The study involved 378 nasopharyngeal samples (UTM® and FLOQSwab™, Copan Diagnostics), including 46 swabs positive for SARS-CoV-2 by RT-PCR. These samples came from asymptomatic (n=99, 26.2%) or symptomatic people (n=279, 73.8%), at different times from symptom onset. The samples were analyzed on LIAISON® XL. Results The overall specificity was 99.4% (CI95% [98.6–100]). The negative predictive value reached 100% in asymptomatic people. Among the 46 positive samples, the overall sensitivity was 84.8% (CI95% [74.4–95.2]), reached 91.9% (CI95% [83.1–100]) in the first fourth days after symptoms onset and was 100% for Cq values ≤25. Antigen was not detected in samples with Cq values >25. Similar results were observed on nasopharyngeal swabs coming from patients infected with the 20I/501Y.V1 variant or the 20H/501Y.V2 variant. Conclusions According to technical performances, the LIAISON® SARS-CoV-2 Ag test may be a useful tool for COVID-19 diagnosis, especially during the first four days of symptoms.

scholarly journals The Performance of Two Rapid Antigen Tests During Population-Level Screening for SARS-CoV-2 Infection

2021 ◽  
Vol 8 ◽  
Author(s):  
Mohammad Alghounaim ◽  
Hamad Bastaki ◽  
Farah Bin Essa ◽  
Hoda Motlagh ◽  
Salman Al-Sabah
Keyword(s):  
Positive Predictive Value ◽  
Prospective Study ◽  
Predictive Value ◽  
Point Of Care ◽  
Population Level ◽  
Lower Sensitivity ◽  
Rt Pcr ◽  
Antigen Tests ◽  
A Prospective Study ◽  
Antigen Testing

Background: SARS-CoV-2 antigen assays offer a rapid mean to diagnose and isolate infected individuals. However, their utility in population-level screening is unknown.Objectives: The performance of two antigen tests in detecting SARS-CoV-2 was assessed among individuals randomly selected in the community.Study Design: A prospective study that performed head-to-head comparison of two SARS-CoV-2 antigen assays. Individuals were recruited during community SARS-CoV-2 screening over 10 working days. Demographic and clinical data were collected. Standard Q COVID-19 Ag test, a point-of-care chromatographic assay, was conducted immediately, and then the sample was transported to the virology laboratory to perform PCR and the LIAISON SARS-CoV-2 Ag chemiluminesence immunoassay.Results: respiratory samples from 991 individuals were collected, and 62 were positive by PCR. Inconclusive PCR results were observed in 19 samples and were excluded. The median age of participants was 40.2 years (IQR 32.3–47.8), and 932 (94%) were males. Most (77.4%) of infections were asymptomatic. The sensitivity and the specificity of the LIAISON assay were 43.3% (95%CI 30.6–56.8) and 99.9% (95%CI 99.3–100). The Standard Q assay had lower sensitivity (30.6%, 95%CI 19.6–43.7) but similar specificity (98.8%, 95%CI, 97.8–99.4). Similarly, the LIAISON assay had higher positive predictive value (96.3%, 95%CI 81–99.9% vs. 63.3%, 95%CI, 43.9–80.1%). Both assays performed better in symptomatic patients and among samples with a low-cycle threshold (Ct < 25).Conclusion: In our setting of random community surveillance, rapid antigen testing of nasopharyngeal swabs by either LIAISON SARS-CoV-2 Ag (DiaSorin) or Standard Q COVID-19 Ag (SD Biosensor) was less sensitive to detecting SARS-CoV-2 than the TaqPath COVID-19 RT-PCR.

scholarly journals High Sensitivity and NPV for BinaxNOW Rapid Antigen Test in Children at a Mass Testing Site During Prevalent Delta Variant

2022 ◽  
Author(s):  
Kristie J Sun ◽  
Mary Jane E Vaeth ◽  
Matthew L Robinson ◽  
Maryam Elhabashy ◽  
Ishaan Gupta ◽  
...  
Keyword(s):  
Predictive Value ◽  
High Sensitivity ◽  
High Volume ◽  
Educational Opportunities ◽  
Antigen Test ◽  
Sample Collection ◽  
Rt Pcr ◽  
Polymerase Chain ◽  
Testing Site ◽  
Antigen Tests

SARS-CoV-2 continues to develop new, increasingly infectious variants, such as delta and omicron. Here, we evaluate the efficacy of the Abbott BinaxNOW Rapid Antigen Test against the gold standard of Reverse Transcription Polymerase Chain Reaction (RT-PCR) in 1054 pediatric participants presenting to a state-owned high-volume Coronavirus Disease 2019 (COVID-19) testing site. During the testing period, the delta variant was predominant. Prior to sample collection, symptomatic and exposure status was collected for all participants based on Centers for Disease Control (CDC) criteria. RT-PCR results demonstrated an overall prevalence rate of 5.2%. For all participants, the sensitivity of the rapid antigen tests was 92.7% (95% CI 82.4% - 98.0%) and specificity was 98.0% (95% CI 97.0%-98.8%). For symptomatic participants, the sensitivity was 92.3% (95% CI 74.9% - 99.1%), specificity was 96.6% (95% CI 93.6%- 98.4%), positive predictive value (PPV) was 72.7% (95% CI 54.5% - 86.7%) and negative predictive value (NPV) was 99.2% (95% CI 98.2% - 100%). Among asymptomatic participants, the sensitivity was 92.6% (95% CI 75.7% - 99.1%), specificity was 98.6% (95% CI 97.5% - 99.3%) the PPV was 71.4% (95% CI 53.7% - 85.4%) and the NPV was 99.7% (95% CI 99.0% - 100%). Our reported sensitivity and NPV are higher than other pediatric studies, but specificity and PPV are lower. Importance Children are especially impacted by the disease and its ability to disrupt educational opportunities. Although vaccinations have been approved for children 5 years and older, many children remain unvaccinated. Widespread testing may improve the ability for children to remain in in-person activities, minimizing absences from school and extracurriculars. Highly accurate rapid antigen tests may be vital to containing future COVID-19 waves while mitigating detrimental effects.

scholarly journals The Value of Rapid Antigen Tests to Identify Carriers of Viable SARS-CoV-2

2021 ◽  
Author(s):  
Elena V. Shidlovskaya ◽  
Nadezhda A. Kuznetsova ◽  
Elizaveta V. Divisenko ◽  
Maria A. Nikiforova ◽  
Andrei E. Siniavin ◽  
...  
Keyword(s):  
Cell Culture ◽  
Diagnostic Value ◽  
Healthy People ◽  
Rt Pcr ◽  
Culture Sensitivity ◽  
Rapid Tests ◽  
Antigen Tests

AbstractThe search for effective methods to detect patients who excrete a viable virus is one of the urgent tasks of modern biomedicine. In the present study, we examined the diagnostic value of two antigen tests BIOCREDIT COVID-19 Ag (RapiGEN Inc., Korea) and SGTI-flex COVID-19 Ag (Sugentech Inc., Korea) for their diagnostic value in identifying patients who excrete viable SARS-CoV-2. As part of the study, we examined samples from 106 patients who had just been admitted to the hospital, who had undergone quantitative RT-PCR and assessment of viability of SARS-CoV-2 using cell culture. Sensitivity was 0.786 (0.492–0.953) for SGTI-flex COVID-19 Ag and 1 (0.768– 1) for Biocredit COVID-19 Ag. Specificity of rapid tests was significantly higher than that of RT-PCR and was 0.663 (0.557–0.758) and 0.674 (0.568–0.768) for SGTI-flex COVID-19 Ag and Biocredit COVID-19 Ag versus 0.304 (0.213–0.409) obtained for PCR. Thus, for tasks of identifying viable SARS-CoV-2 during screening of conditionally healthy people, as well as monitoring those quarantined, rapid tests show significantly better results.

scholarly journals Sensitivity of nasopharyngeal, oropharyngeal and nasal washes specimens for SARS-CoV-2 detection in the setting of sampling device shortage

2020 ◽  
Author(s):  
Adrien Calame ◽  
Lena Mazza ◽  
Adriana Renzoni ◽  
Laurent Kaiser ◽  
Manuel Schibler
Keyword(s):  
Culture Medium ◽  
Alternative Methods ◽  
Analytical Sensitivity ◽  
Sample Collection ◽  
Rt Pcr ◽  
Sampling Device ◽  
Viral Transport ◽  
Viral Transport Medium ◽  
A Cell ◽  
Oropharyngeal Swab

In the context of an unprecedented shortage of nasopharyngeal swabs (NPS) or sample transport media during the coronavirus disease 2019 (COVID-19) crisis, alternative methods for sample collection are needed. To address this need, we validated a cell culture medium as a viral transport medium, and compared the analytical sensitivity of SARS-CoV-2 real-time RT-PCR in nasal wash (NW), oropharyngeal swab (OPS) and NPS specimens. Both the clinical and analytical sensitivity were comparable in these three sample types. OPS and NW specimens may therefore represent suitable alternatives to NPS for SARS-CoV-2 detection.

scholarly journals The evaluation of a novel digital immunochromatographic assay with silver amplification to detect SARS-CoV-2

2021 ◽  
Author(s):  
Yoko Kurihara ◽  
Yoshihiko Kiyasu ◽  
Yusaku Akashi ◽  
Yuto Takeuchi ◽  
Kenji Narahara ◽  
...  
Keyword(s):  
Predictive Value ◽  
Limit Of Detection ◽  
The Other ◽  
Nucleic Acid Amplification ◽  
Immunochromatographic Assay ◽  
Clinical Samples ◽  
Antigen Test ◽  
Rt Pcr ◽  
Reverse Transcription Pcr ◽  
Antigen Tests

Introduction Rapid antigen tests are convenient for diagnosing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2); however, they have lower sensitivities than nucleic acid amplification tests. In this study, we evaluated the diagnostic performance of Quick Chaser Auto SARS-CoV-2, a novel digital immunochromatographic assay that is expected to have higher sensitivity than conventional antigen tests. Methods A prospective observational study was conducted between February 8 and March 24, 2021. We simultaneously obtained two nasopharyngeal samples, one for evaluation with the QuickChaser Auto SARS-CoV-2 antigen test and the other for assessment with reverse transcription PCR (RT-PCR), considered the gold-standard reference test. The limit of detection (LOD) of the new antigen test was compared with those of four other commercially available rapid antigen tests. Results A total of 1401 samples were analyzed. SARS-CoV-2 was detected by reference RT-PCR in 83 (5.9%) samples, of which 36 (43.4%) were collected from symptomatic patients. The sensitivity, specificity, positive predictive value, and negative predictive value were 74.7% (95% confidence interval (CI): 64.0-83.6%), 99.8% (95% CI: 99.5-100%), 96.9% (95% CI: 89.2-99.6%), and 98.4% (95% CI: 97.6-99.0%), respectively. When limited to samples with a cycle threshold (Ct) <30 or those from symptomatic patients, the sensitivity increased to 98.3% and 88.9%, respectively. The QuickChaser Auto SARS-CoV-2 detected 34-120 copies/test, which indicated greater sensitivity than the other rapid antigen tests. Conclusions QuickChaser Auto SARS-CoV-2 showed sufficient sensitivity and specificity in clinical samples of symptomatic patients. The sensitivity was comparable to RT-PCR in samples with Ct<30.

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