The impact of Sample Processing on the Rapid Antigen Detection test for SARS-CoV-2: Virus Inactivation, VTM Selection, and Sample Preservation.
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ABSTRACT:
SUBMITTER: Zhou H
PROVIDER: S-EPMC8426133 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the causative agent of COVID-19 disease. RT-qPCR has been the primary method of diagnosis; however, the required infrastructure is lacking in many developing countries and the virus has remained a global challenge. More inexpensive and immediate test methods are required to facilitate local, regional, and national management strategies to re-open world economies. Here we have developed a SARS-CoV-2 antigen test in an inexpensive lateral flow format to generate a chromatographic result identifying the presence of the SARS-CoV-2 antigen, and thus an active infection, within a patient anterior nares swab sample. Our 15-min test requires no equipment or laboratory infrastructure to administer with a limit of detection of 2.0 × 102 TCID50/mL and 87.5% sensitivity, 100% specificity when tested against 40 known positive and 40 known negative patient samples established by a validated RT-qPCR test.
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Project description:COVID-19 diagnostics was quickly ramped up worldwide early 2020 based on the detection of viral RNA. However, based on the scientific knowledge for pre-existing coronaviruses, it was expected that the SARS-CoV-2 RNA will be detected from symptomatic and at significant rates also from asymptomatic individuals due to persistence of non-infectious RNA. To increase the efficacy of diagnostics, surveillance, screening and pandemic control, rapid methods, such as antigen tests, are needed for decentralized testing and to assess infectiousness. A novel automated mariPOC SARS-CoV-2 test was developed for the detection of conserved structural viral nucleocapsid proteins. The test utilizes sophisticated optical laser technology for two-photon excitation and individual detection of immunoassay solid-phase particles. We validated the new method against qRT-PCR. Sensitivity of the test was 100.0% (13/13) directly from nasopharyngeal swab specimens and 84.4% (38/45) from swab specimens in undefined transport mediums. Specificity of the test was 100.0% (201/201). The test's limit of detection was 2.7 TCID50/test. It showed no cross-reactions. Our study shows that the new test can detect infectious individuals already in 20 min with clinical sensitivity close to qRT-PCR. The mariPOC is a versatile platform for syndromic testing and for high capacity infection control screening of infectious individuals.
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Project description:Lateral flow immunochromatographic rapid diagnostic tests (RDTs) are the primary form of medical diagnostic used for malaria in underdeveloped nations. Unfortunately, many of these tests do not detect asymptomatic malaria carriers. In order for eradication of the disease to be achieved, this problem must be solved. In this study, we demonstrate enhancement in the performance of six RDT brands when a simple sample-processing step is added to the front of the diagnostic process. Greater than a 4-fold RDT signal enhancement was observed as a result of the sample processing step. This lowered the limit of detection for RDT brands to submicroscopic parasitemias. For the best performing RDTs the limits of detection were found to be as low as 3 parasites per ?L. Finally, through individual donor samples, the correlations between donor source, WHO panel detection scores and RDT signal intensities were explored.
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Project description:ObjectivesQuick and inexpensive SARS-CoV-2 screening and frontline testing are in growing demand. Our study aimed to evaluate the performance of the immunochromatographic AMP rapid antigen test (AMP RAT) compared to the gold-standard real-time reverse transcription PCR (rRT-PCR) in a hospital cohort.MethodsA total of 392 patients, who presented consecutively with COVID-19 symptoms in our emergency department, were included in this retrospective study. Two swabs were collected per patient: a nasopharyngeal for the RAT and a combined naso- and oropharyngeal for the rRT-PCR. A positive rRT-PCR (defined as cycle threshold (Ct) < 40) was found in 94 (24%) patients.ResultsIn our cohort with a median patient age of 70, overall sensitivity and specificity of the AMP RAT was 69.2% (58.8-78.3, 95% CI) and 99.7% (98.1-100.0, 95% CI), respectively. In patients with a Ct value < 25 and < 30, higher sensitivities of 100.0% (89.4-100.0, 95% CI) and 91.8% (81.9-97.3%, 95% CI) were observed.ConclusionsThe AMP RAT showed a high sensitivity in patients with a Ct value < 25 and < 30 and might be helpful for frontline testing whenever rRT-PCR is not readily available.
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Project description:COVID-19 has evolved into a global pandemic. Early and rapid detection is crucial to control of the SARS-CoV-2 transmission. While representing the gold standard for early diagnosis, nucleic acid tests for SARS-CoV-2 are often complicated and time-consuming. Serological rapid antibody tests are characterized by high rates of false-negative diagnoses, especially during early infection. Here, we developed a novel nanozyme-based chemiluminescence paper assay for rapid and sensitive detection of SARS-CoV-2 spike antigen, which integrates nanozyme and enzymatic chemiluminescence immunoassay with the lateral flow strip. The core of our paper test is a robust Co-Fe@hemin-peroxidase nanozyme that catalyzes chemiluminescence comparable with natural peroxidase HRP and thus amplifies immune reaction signal. The detection limit for recombinant spike antigen of SARS-CoV-2 was 0.1 ng/mL, with a linear range of 0.2-100 ng/mL. Moreover, the sensitivity of test for pseudovirus could reach 360 TCID50/mL, which was comparable with ELISA method. The strip recognized SARS-CoV-2 antigen specifically, and there was no cross reaction with other coronaviruses or influenza A subtypes. This testing can be completed within 16 min, much shorter compared to the usual 1-2 h required for currently used nucleic acid tests. Furthermore, signal detection is feasible using the camera of a standard smartphone. Ingredients for nanozyme synthesis are simple and readily available, considerably lowering the overall cost. In conclusion, our paper test provides a high-sensitive point-of-care testing (POCT) approach for SARS-CoV-2 antigen detection, which should greatly facilitate early screening of SARS-CoV-2 infections, and considerably lower the financial burden on national healthcare resources.
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Project description:Airborne transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is one of the leading mechanisms of spread, especially in confined environments. The study aims to assess the thermal inactivation of SARS-CoV-2 at high temperatures in the time scale of seconds. An electric heater with a coiled resistance wire is located perpendicularly to the airflow direction inside an air tunnel. The airflow rate through the tunnel was 0.6 m3/h (10 L/ min). SARS-CoV-2 were suspended in Dulbecco's modified Eagle's medium (DMEM) with 10 % fetal bovine serum (FBS), aerosolized by a nebulizer at a rate of 0.2 L/min and introduced to the airflow inside the heater with the use of a compressor and an aspirator. In the control experiment, with the heater off, SARS-CoV-2 passed through the system. In the virus inactivation test experiments, the heater's outlet air temperature was set to 150 ± 5 °C and 220 ± 5 °C, and the air traveling through the tunnel was exposed to heat for 1.44 s. An inline gelatine filter harvested SARS-CoV-2 that passed through the system. The viral titer obtained from the gelatine filter in the control experiment was about 5.5 log10 TCID50. The virus's loss in viability in test experiments at 150 °C and 220 °C were 99.900 % and 99.999 %, respectively. The results indicate that high-temperature thermal inactivation substantially reduces the concentration of SARS-CoV-2 in the air within seconds.
| S-EPMC8757645 | biostudies-literature
Project description:The current severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has had an enormous impact on society worldwide, threatening the lives and livelihoods of many. The effects will continue to grow and worsen if economies begin to open without the proper precautions, including expanded diagnostic capabilities. To address this need for increased testing, we have developed a sensitive reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay compatible with current reagents, which utilizes a colorimetric readout in as little as 30 min. A rapid inactivation protocol capable of inactivating virions, as well as endogenous nucleases, was optimized to increase sensitivity and sample stability. This protocol, combined with the RT-LAMP assay, has a sensitivity of at least 50 viral RNA copies per microliter in a sample. To further increase the sensitivity, a purification protocol compatible with this inactivation method was developed. The inactivation and purification protocol, combined with the RT-LAMP assay, brings the sensitivity to at least 1 viral RNA copy per microliter in a sample. This simple inactivation and purification pipeline is inexpensive and compatible with other downstream RNA detection platforms and uses readily available reagents. It should increase the availability of SARS-CoV-2 testing as well as expand the settings in which this testing can be performed.
| S-EPMC7533677 | biostudies-literature
Project description:BackgroundSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causing coronavirus disease 2019 (COVID-19) is currently finally determined in laboratory settings by real-time reverse-transcription polymerase-chain-reaction (rt-PCR). However, simple testing with immediately available results are crucial to gain control over COVID-19. The aim was to evaluate such a point-of-care antigen rapid test (AG-rt) device in its performance compared to laboratory-based rt-PCR testing in COVID-19 suspected, symptomatic patients.MethodsFor this prospective study, two specimens each of 541 symptomatic female (54.7%) and male (45.3%) patients aged between 18 and 95 years tested at five emergency departments (ED, n = 296) and four primary healthcare centres (PHC, n = 245), were compared, using AG-rt (positive/negative/invalid) and rt-PCR (positive/negative and cycle threshold, Ct) to diagnose SARS-CoV-2. Diagnostic accuracy, sensitivity, specificity, positive predictive values (PPV), negative predictive value (NPV), and likelihood ratios (LR+/-) of the AG-rt were assessed.ResultsDifferences between ED and PHC were detected regarding gender, age, symptoms, disease prevalence, and diagnostic performance. Overall, 174 (32.2%) were tested positive on AG-rt and 213 (39.4%) on rt-PCR. AG correctly classified 91.7% of all rt-PCR positive cases with a sensitivity of 80.3%, specificity of 99.1%, PPV of 98.3, NPV of 88.6%, LR(+) of 87.8, and LR(-) of 0.20. The highest sensitivities and specificities of AG-rt were detected in PHC (sensitivity: 84.4%, specificity: 100.0%), when using Ct of 30 as cut-off (sensitivity: 92.5%, specificity: 97.8%), and when symptom onset was within the first three days (sensitivity: 82.9%, specificity: 99.6%).ConclusionsThe highest sensitivity was detected with a high viral load. Our findings suggest that AG-rt are comparable to rt-PCR to diagnose SARS-CoV-2 in COVID-19 suspected symptomatic patients presenting both at emergency departments and primary health care centres.
| S-EPMC8629250 | biostudies-literature
Project description:BackgroundSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can spread from symptomatic patients with COVID-19, but also from asymptomatic individuals. Therefore, robust surveillance and timely interventions are essential for the control of virus spread within the community. In this regard the frequency of testing and speed of reporting, but not the test sensitivity alone, play a crucial role.ObjectivesIn order to reduce the costs and meet the expanding demands in real-time RT-PCR (rRT-PCR) testing for SARS-CoV-2, complementary assays, such as rapid antigen tests, have been developed. Rigorous analysis under varying conditions is required to assess the clinical performance of these tests and to ensure reproducible results.ResultsWe evaluated the sensitivity and specificity of a recently licensed rapid antigen test using 137 clinical samples in two institutions. Test sensitivity was between 88.2-89.6 % when applied to samples with viral loads typically seen in infectious patients. Of 32 rRT-PCR positive samples, 19 demonstrated infectivity in cell culture, and 84 % of these samples were reactive with the antigen test. Seven full-genome sequenced SARS-CoV-2 isolates and SARS-CoV-1 were detected with this antigen test, with no cross-reactivity against other common respiratory viruses.ConclusionsNumerous antigen tests are available for SARS-CoV-2 testing and their performance to detect infectious individuals may vary. Head-to-head comparison along with cell culture testing for infectivity may prove useful to identify better performing antigen tests. The antigen test analyzed in this study is easy-to-use, inexpensive, and scalable. It can be helpful in monitoring infection trends and thus has potential to reduce transmission.
| S-EPMC7832367 | biostudies-literature