scholarly journals Quantifying SARS-CoV-2 nucleocapsid antigen in oropharyngeal swabs using single molecule array technology

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Dorte Aa. Olsen ◽  
Claus L. Brasen ◽  
Søren Kahns ◽  
Jeppe B. Madsen ◽  
Helene Kierkegaard ◽  
...  
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Point Of Care ◽  
Reference Method ◽  
Rt Pcr ◽  
Clinical Sensitivity ◽  
Rna Molecules ◽  
Antigen Assay ◽  
Array Technology ◽  
Highly Sensitive

AbstractThis study aimed to develop a highly sensitive SARS-CoV-2 nucleocapsid antigen assay using the single molecule array (Simoa) technology and compare it with real time RT-PCR as used in routine clinical practice with the ambition to achieve a comparative technical and clinical sensitivity. Samples were available from 148 SARS-CoV-2 real time RT-PCR positive and 73 SARS-CoV-2 real time RT-PCR negative oropharyngeal swabs. For determination of technical sensitivity SARS-CoV-2 virus culture material was used. The samples were treated with lysis buffer and analyzed using both an in-house and a pre-commercial SARS-CoV-2 nucleocapsid antigen assay on Simoa. Both nucleocapsid antigen assays have a technical sensitivity corresponding to around 100 SARS-CoV-2 RNA molecules/mL. Using a cut-off at 0.1 pg/mL the pre-commercial SARS-CoV-2 nucleocapsid antigen assay had a sensitivity of 96% (95% CI 91.4–98.5%) and specificity of 100% (95% CI 95.1–100%). In comparison the in-house nucleocapsid antigen assay had sensitivity of 95% (95% CI 89.3–98.1%) and a specificity of 100% (95% CI 95.1–100%) using a cut-off at 0.01 pg/mL. The two SARS-CoV-2 nucleocapsid antigen assays correlated with r = 0.91 (P < 0.0001). The in-house and the pre-commercial SARS-CoV-2 nucleocapsid antigen assay demonstrated technical and clinical sensitivity comparable to real-time RT-PCR methods for identifying SARS-CoV-2 infected patients and thus can be used clinically as well as serve as a reference method for antigen Point of Care Testing.

scholarly journals Quantifying SARS-Cov-2 Nucleocapsid Antigen in Oropharyngeal Swabs Using Single Molecule Array Technology

2021 ◽  
Author(s):  
Dorte Aa. Olsen ◽  
Claus L. Brasen ◽  
Søren Kahns ◽  
Jeppe B. Madsen ◽  
Helene Kierkegaard ◽  
...  
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Point Of Care ◽  
Reference Method ◽  
Rt Pcr ◽  
Clinical Sensitivity ◽  
Rna Molecules ◽  
Antigen Assay ◽  
Array Technology ◽  
Highly Sensitive

Abstract Background: This study aimed to develop a highly sensitive SARS-CoV-2 nucleocapsid antigen assay using the single molecule array (Simoa) technology and compare it with real time RT-PCR as used in routine clinical practice with the ambition to achieve a comparative technical and clinical sensitivity. Methods: Samples were available from 148 SARS-CoV-2 real time RT-PCR positive and 73 SARS-CoV-2 real time RT-PCR negative oropharyngeal swabs. For determination of technical sensitivity SARS-CoV-2 virus culture material was used. The samples were treated with lysis buffer and analyzed using both an in-house and a pre-commercial SARS-CoV-2 nucleocapsid antigen assay on Simoa. Results: Both nucleocapsid antigen assays have a technical sensitivity corresponding to around 100 SARS-CoV-2 RNA molecules/mL. Using a cut-off at 0.1 pg/mL the pre-commercial SARS-CoV-2 nucleocapsid antigen assay had a sensitivity of 96% (95% CI, 91.4-98.5%) and specificity of 100% (95% CI, 95.1-100%). In comparison the in-house nucleocapsid antigen assay had sensitivity of 95% (95% CI, 89.3-98.1%) and a specificity of 100% (95% CI, 95.1-100%) using a cut-off at 0.01 pg/mL. The two SARS-CoV-2 nucleocapsid antigen assays correlated with r=0.91 (p<0.0001). Conclusions: The in-house and the pre-commercial SARS-CoV-2 nucleocapsid antigen assay demonstrated technical and clinical sensitivity comparable to real-time RT-PCR methods for identifying SARS-CoV-2 infected patients and thus can be used clinically as well as serve as a reference method for antigen Point of Care Testing.

Highly Sensitive Detection of Paraquat with Pillar[5]arene as an aptamer in α-Hemolysin Nanopore

2021 ◽  
Author(s):  
Xiaojia Jiang ◽  
Mingsong Zang ◽  
Fei Li ◽  
Chunxi Hou ◽  
Quan Luo ◽  
...  
Keyword(s):  
Real Time ◽  
High Throughput ◽  
Single Molecule ◽  
Single Molecule Detection ◽  
Sensitive Detection ◽  
High Throughput Analysis ◽  
Throughput Analysis ◽  
The Real ◽  
Highly Sensitive ◽  
Highly Sensitive Detection

Biological nanopore-based techniques have attracted more and more attention recently in the field of single-molecule detection, because they allow the real-time, sensitive, high-throughput analysis. Herein, we report an engineered biological...

scholarly journals Evaluation of mobile real-time polymerase chain reaction tests for the detection of severe acute respiratory syndrome coronavirus 2

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Chukwunonso Onyilagha ◽  
Henna Mistry ◽  
Peter Marszal ◽  
Mathieu Pinette ◽  
Darwyn Kobasa ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Real Time ◽  
Point Of Care ◽  
Middle Eastern ◽  
Turnaround Time ◽  
Cross Reactivity ◽  
Clinical Samples ◽  
Diarrhea Virus ◽  
Highly Sensitive ◽  
Transmissible Gastroenteritis

AbstractThe coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), calls for prompt and accurate diagnosis and rapid turnaround time for test results to limit transmission. Here, we evaluated two independent molecular assays, the Biomeme SARS-CoV-2 test, and the Precision Biomonitoring TripleLock SARS-CoV-2 test on a field-deployable point-of-care real-time PCR instrument, Franklin three9, in combination with Biomeme M1 Sample Prep Cartridge Kit for RNA 2.0 (M1) manual extraction system for rapid, specific, and sensitive detection of SARS-COV-2 in cell culture, human, and animal clinical samples. The Biomeme SARS-CoV-2 assay, which simultaneously detects two viral targets, the orf1ab and S genes, and the Precision Biomonitoring TripleLock SARS-CoV-2 assay that targets the 5′ untranslated region (5′ UTR) and the envelope (E) gene of SARS-CoV-2 were highly sensitive and detected as low as 15 SARS-CoV-2 genome copies per reaction. In addition, the two assays were specific and showed no cross-reactivity with Middle Eastern respiratory syndrome coronavirus (MERS-CoV), infectious bronchitis virus (IBV), porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis (TGE) virus, and other common human respiratory viruses and bacterial pathogens. Also, both assays were highly reproducible across different operators and instruments. When used to test animal samples, both assays equally detected SARS-CoV-2 genetic materials in the swabs from SARS-CoV-2-infected hamsters. The M1 lysis buffer completely inactivated SARS-CoV-2 within 10 min at room temperature enabling safe handling of clinical samples. Collectively, these results show that the Biomeme and Precision Biomonitoring TripleLock SARS-CoV-2 mobile testing platforms could reliably and promptly detect SARS-CoV-2 in both human and animal clinical samples in approximately an hour and can be used in remote areas or health care settings not traditionally serviced by a microbiology laboratory.

A novel and highly sensitive real-time nested RT-PCR assay in a single closed tube for detection of enterovirus

2018 ◽  
Vol 90 (3) ◽  
pp. 181-185 ◽  
Author(s):  
Xin-xin Shen ◽  
Fang-zhou Qiu ◽  
Huai-long Zhao ◽  
Meng-jie Yang ◽  
Liu Hong ◽  
...  
Keyword(s):  
Real Time ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Highly Sensitive ◽  
Closed Tube

scholarly journals Development and Evaluation of Novel and Highly Sensitive Single-Tube Nested Real-Time RT-PCR Assays for SARS-CoV-2 Detection

2020 ◽  
Vol 21 (16) ◽  
pp. 5674
Author(s):  
Cyril Chik-Yan Yip ◽  
Siddharth Sridhar ◽  
Kit-Hang Leung ◽  
Anthony Chin-Ki Ng ◽  
Kwok-Hung Chan ◽  
...  
Keyword(s):  
Real Time ◽  
Limit Of Detection ◽  
Rt Pcr ◽  
Single Tube ◽  
Highly Sensitive ◽  
Testing Algorithms ◽  
Pcr Assays ◽  
Human Coronaviruses ◽  
Pooled Sample

Sensitive molecular assays are critical for coronavirus disease 2019 (COVID-19) diagnosis. Here, we designed and evaluated two single-tube nested (STN) real-time RT-PCR assays, targeting SARS-CoV-2 RdRp/Hel and N genes. Both STN assays had a low limit of detection and did not cross react with other human coronaviruses and respiratory viruses. Using 213 initial respiratory specimens from suspected COVID-19 patients, the sensitivity of both the STN COVID-19-RdRp/Hel and the STN COVID-19-N assays was 100% (99/99), while that of the comparator non-nested N assay was 95% (94/99). Among 108 follow-up specimens from confirmed COVID-19 patients who tested negative by the non-nested COVID-19-RdRp/Hel assay, 28 (25.9%) were positive for SARS-CoV-2 by the STN COVID-19-RdRp/Hel or the STN COVID-19-N assay. To evaluate the performance of our novel STN assays in pooled specimens, we created four sample pools, with each pool consisting of one low positive specimen and 49 negative specimens. While the non-nested COVID-19-RdRp/Hel assay was positive in only one of four sample pools (25%), both of the STN assays were positive in two of four samples pools (50%). In conclusion, the STN assays are highly sensitive and specific for SARS-CoV-2 detection. Their boosted sensitivity offers advantages in non-traditional COVID-19 testing algorithms such as saliva screening and pooled sample screening during massive screening.

scholarly journals The efficiency of sodC gene / N. meningitidis detection in comparison with the classical methods for the diagnosis of meningococcal infection / Evaluarea eficienţei Real Time PCR TaqMan utilizând gena sodC / N. meningitidis în comparaţie cu metodele clasice utilizate în diagnosticul infecţiei meningococice

2015 ◽  
Vol 23 (1) ◽  
Author(s):  
Roxana Elena Nemescu ◽  
Ramona Gabriela Ursu ◽  
Carmen Mihaela Dorobăț ◽  
Luminița Smaranda Iancu
Keyword(s):  
Antibiotic Therapy ◽  
Real Time ◽  
Real Time Pcr ◽  
Latex Agglutination ◽  
Blood Analysis ◽  
Meningococcal Infection ◽  
Specific Method ◽  
Rt Pcr ◽  
Highly Sensitive ◽  
Etiological Agents

AbstractMeningococcal infection requires a fast and accurate diagnostic method in order to correctly initiate the antibiotic therapy. The aim of our study was to assess the efficiency of Real Time PCR -Taq Man using sod C gene / N. meningitidis in comparison with the classical methods for the diagnosis of meningococcal infection - direct microscopy, cultivation, latex agglutination and blood culture. We have detected 24/44 (54.54%) patients with meningococcal infection. In both cases of patients with / without previous antibiotic therapy before admission, the AUC (area under curve) had the highest values for RT PCR in CSF and blood analysis. This sod C RT-PCR assay is a highly sensitive and specific method for detection of Neisseria meningitis and it would be useful to include this method like a multiplex in routine testing of patients with clinical meningococcal infection for other etiological agents also.

Comparative Evaluation of a Phage Protein Ligand Assay with Real-Time PCR and a Reference Method for the Detection of Escherichia coli O157:H7 in Raw Ground Beef and Trimmings

2011 ◽  
Vol 74 (1) ◽  
pp. 6-12 ◽  
Author(s):  
F. SAVOYE ◽  
P. FENG ◽  
C. ROZAND ◽  
M. BOUVIER ◽  
A. GLEIZAL ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Real Time ◽  
Ground Beef ◽  
Reference Method ◽  
Enterohemorrhagic Escherichia Coli ◽  
Detection Methods ◽  
Escherichia Coli O157 ◽  
Rt Pcr ◽  
Raw Meat ◽  
E Coli

Enterohemorrhagic Escherichia coli O157:H7 is an important pathogen associated with infections caused by consumption of undercooked raw meat. Sensitive and rapid detection methods for E. coli O157:H7 are essential for the meat industry to ensure a safe meat supply. This study was conducted to compare the sensitivity of the VIDAS ultraperformance E. coli test (ECPT UP) with a noncommercial real-time (RT) PCR method and the U.S. Department of Agriculture, Food Safety and Inspection Service (USDA-FSIS) reference method for detecting E. coli O157:H7 in raw ground beef. Optimal enrichment times and the efficacy of testing different types of raw meat, either as individual samples (25 g) or as composites (375 g), were examined. For 25-g samples of each type of raw ground beef tested, 6 h of enrichment was sufficient for both the VIDAS ECPT UP and RT-PCR methods, but for 375-g samples, 24 h of enrichment was required. Both the VIDAS ECPT UP and RT-PCR methods produced results similar to those obtained with the USDA-FSIS reference method after 18 to 24 h of enrichment. The primer specificity of the RT-PCR assay and the highly specific phage ligand used in the VIDAS ECPT UP for target recognition enabled the detection of low levels of E. coli O157:H7 in 25 g of various types of raw ground beef. The tests also allowed the detection of E. coli O157:H7 in composite raw ground beef and trimmings in samples of up to 375 g.

scholarly journals Development, Evaluation of the PNA RT-LAMP Assay for Rapid Molecular Detection of SARS-CoV-2

2021 ◽  
Author(s):  
Chinbayar Bat-Ochir ◽  
Yeon-Sook Kim ◽  
Han Gyeul Kim ◽  
See Sok Lee ◽  
Han Woo Lee ◽  
...  
Keyword(s):  
Real Time ◽  
Lamp Assay ◽  
Study Data ◽  
Molecular Diagnostic ◽  
Nasopharyngeal Swab ◽  
Rt Pcr ◽  
Clinical Sensitivity ◽  
Clinical Accuracy ◽  
Absolute Detection Limit ◽  
Development Evaluation

Abstract Dual-labeled PNA probe used RT-LAMP molecular rapid assay targeting SARS-CoV-2 ORF1ab and N genes was developed, and the analytical, clinical performances for detection of SARS-CoV-2 RNA extracted from clinical nasopharyngeal swab specimens were evaluated in this study. Data showed that this assay is highly specific for SARS-CoV-2, and the absolute detection limit is 1 genomic copy per microliter of viral RNA which can be considered to be comparable to gold-standard molecular diagnostic method real-time reverse transcriptase PCR. Both clinical sensitivity and specificity against a commercial real-time RT-PCR assay were determined as identical. In conclusion, the PNA RT-LAMP assay showed high analytical and clinical accuracy which are identical to real-time RT-PCR which has been routinely used for the detection of SARS-CoV-2.

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