Comparison of RT-PCR, RT-LAMP, and Antigen Quantification Assays for the Detection of SARS-CoV-2
A rapid and simple alternative test to real-time reverse transcription-polymerase chain reaction (RT-PCR) is required for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to help curb the spread of coronavirus disease (COVID-19). In the present study, we compared the RT-PCR method with c...
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| Published in | Japanese Journal of Infectious Diseases Vol. 75; no. 3; pp. 249 - 253 |
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| Main Authors | , , , , , |
| Format | Journal Article |
| Language | English |
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Japan
National Institute of Infectious Diseases, Japanese Journal of Infectious Diseases Editorial Committee
31.05.2022
Japan Science and Technology Agency |
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| Abstract | A rapid and simple alternative test to real-time reverse transcription-polymerase chain reaction (RT-PCR) is required for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to help curb the spread of coronavirus disease (COVID-19). In the present study, we compared the RT-PCR method with chemiluminescent enzyme immunoassay (CLEIA) and reverse transcription loop-mediated isothermal amplification (RT-LAMP). We observed that the number of SARS-CoV-2 RNA copies and the CLEIA antigen quantification values were highly correlated. The detection limit for antigen quantification was 42.8 RNA copies for saliva samples and 23.4 copies for nasopharyngeal swab samples. For both purified RNA and purification-free crude RNA, the number of RNA copies and RT-LAMP threshold time (Tt) values were inversely correlated. RT-LAMP with purified RNA detected low copy numbers of RNA (5–50 copies), whereas fewer than 250 RNA copies could not be detected using crude RNA. CLEIA antigen quantification is potentially useful for large-scale screening, as it is compatible with high-throughput testing. RT-LAMP with crude RNA samples is applicable for rapid point-of-care testing because it can directly use patient specimens. It is important to select a diagnostic method that is simple and rapid when compared with RT-PCR, depending on the situation. |
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| AbstractList | A rapid and simple alternative test to real-time reverse transcription-polymerase chain reaction (RT-PCR) is required for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to help curb the spread of coronavirus disease (COVID-19). In the present study, we compared the RT-PCR method with chemiluminescent enzyme immunoassay (CLEIA) and reverse transcription loop-mediated isothermal amplification (RT-LAMP). We observed that the number of SARS-CoV-2 RNA copies and the CLEIA antigen quantification values were highly correlated. The detection limit for antigen quantification was 42.8 RNA copies for saliva samples and 23.4 copies for nasopharyngeal swab samples. For both purified RNA and purification-free crude RNA, the number of RNA copies and RT-LAMP threshold time (Tt) values were inversely correlated. RT-LAMP with purified RNA detected low copy numbers of RNA (5-50 copies), whereas fewer than 250 RNA copies could not be detected using crude RNA. CLEIA antigen quantification is potentially useful for large-scale screening, as it is compatible with high-throughput testing. RT-LAMP with crude RNA samples is applicable for rapid point-of-care testing because it can directly use patient specimens. It is important to select a diagnostic method that is simple and rapid when compared with RT-PCR, depending on the situation. A rapid and simple alternative test to real-time reverse transcription-polymerase chain reaction (RT-PCR) is required for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to help curb the spread of coronavirus disease (COVID-19). In the present study, we compared the RT-PCR method with chemiluminescent enzyme immunoassay (CLEIA) and reverse transcription loop-mediated isothermal amplification (RT-LAMP). We observed that the number of SARS-CoV-2 RNA copies and the CLEIA antigen quantification values were highly correlated. The detection limit for antigen quantification was 42.8 RNA copies for saliva samples and 23.4 copies for nasopharyngeal swab samples. For both purified RNA and purification-free crude RNA, the number of RNA copies and RT-LAMP threshold time (Tt) values were inversely correlated. RT-LAMP with purified RNA detected low copy numbers of RNA (5-50 copies), whereas fewer than 250 RNA copies could not be detected using crude RNA. CLEIA antigen quantification is potentially useful for large-scale screening, as it is compatible with high-throughput testing. RT-LAMP with crude RNA samples is applicable for rapid point-of-care testing because it can directly use patient specimens. It is important to select a diagnostic method that is simple and rapid when compared with RT-PCR, depending on the situation.A rapid and simple alternative test to real-time reverse transcription-polymerase chain reaction (RT-PCR) is required for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to help curb the spread of coronavirus disease (COVID-19). In the present study, we compared the RT-PCR method with chemiluminescent enzyme immunoassay (CLEIA) and reverse transcription loop-mediated isothermal amplification (RT-LAMP). We observed that the number of SARS-CoV-2 RNA copies and the CLEIA antigen quantification values were highly correlated. The detection limit for antigen quantification was 42.8 RNA copies for saliva samples and 23.4 copies for nasopharyngeal swab samples. For both purified RNA and purification-free crude RNA, the number of RNA copies and RT-LAMP threshold time (Tt) values were inversely correlated. RT-LAMP with purified RNA detected low copy numbers of RNA (5-50 copies), whereas fewer than 250 RNA copies could not be detected using crude RNA. CLEIA antigen quantification is potentially useful for large-scale screening, as it is compatible with high-throughput testing. RT-LAMP with crude RNA samples is applicable for rapid point-of-care testing because it can directly use patient specimens. It is important to select a diagnostic method that is simple and rapid when compared with RT-PCR, depending on the situation. |
| ArticleNumber | JJID.2021.476 |
| Author | Tanimoto, Yoshihiko Miyamoto, Sonoko Arikawa, Kentaro Iwamoto, Tomotada Mori, Ai Ito, Erika |
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| Cites_doi | 10.1093/cid/ciaa1388 10.1016/j.cca.2021.02.014 10.1093/nar/28.12.e63 10.1136/bmj.m3862 10.1016/j.jcv.2021.104764 10.1016/j.ijid.2020.08.029 10.1128/JCM.00726-13 10.1136/bmjresp-2020-000830 10.1038/s41586-020-2196-x 10.20965/jdr.2021.p0084 10.1016/j.jinf.2021.04.007 10.1016/S0140-6736(20)30183-5 10.1016/j.jviromet.2006.11.031 10.7883/yoken.JJID.2020.061 |
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| References_xml | – reference: 1. Huang C, Wang Y, Li X, et al. Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China. Lancet. 2020; 395:497-506. – reference: 3. Shirato K, Nao N, Katano H, et al. Development of genetic diagnostic methods for detection for novel coronavirus 2019(nCoV-2019) in Japan. Jpn J Infect Dis. 2020;73:304-307. – reference: 4. Shinkai N, Matsuura K, Sugauchi F, et al. Application of a newly developed high-sensitivity HBsAg chemiluminescent enzyme immunoassay for hepatitis B patients with HBsAg seroclearance. J Clin Microbiol. 2013;51:3484-3491. – reference: 9. Hirotsu Y, Maejima M, Shibusawa M, et al. Comparison of automated SARS-CoV-2 antigen test for COVID-19 infection with quantitative RT-PCR using 313 nasopharyngeal swabs, including from seven serially followed patients. Int J Infect Dis. 2020;99:397-402. – reference: 8. Florkowski CM. Sensitivity, specificity, receiver-operating characteristic (ROC) curves and likelihood ratios: communicating the performance of diagnostic tests. Clin Biochem Rev. 2008;29(Suppl 1):S83-S87. – reference: 5. Notomi T, Okayama H, Masubuchi H, et al. Loop-mediated isothermal amplification of DNA. Nucleic Acids Res. 2000;28:e63. – reference: 7. Basso D, Aita A, Padoan A, et al. Salivary SARS-CoV-2 antigen rapid detection: A prospective cohort study. Clin Chim Acta. 2021;517:54-59. – reference: 6. Kurosaki Y, Takada A, Ebihara H, et al. Rapid and simple detection of Ebola virus by reverse transcription-loop-mediated isothermal amplification. J Virol Methods. 2007;141:78-83. – reference: 2. Nakanishi N, Iijima Y. The novel coronavirus pandemic and the state of the epidemic in Kobe, Japan. J Disaster Res. 2021;16:84-87. – reference: 13. Yokota I, Shane PY, Okada K, et al. Mass screening of asymptomatic persons for severe acute respiratory syndrome coronavirus 2 using saliva. Clin Infect Dis. 2021;73:e559-e565. – reference: 12. Kodana M, Kitagawa Y, Takahashi R, et al. Concerns about the clinical usefulness of saliva specimens for the diagnosis of COVID-19. J Infect. 2021;83:119-145. – reference: 11. Cevik M, Kuppalli K, Kindrachuk J, et al. Virology, transmission, and pathogenesis of SARS-CoV-2. BMJ. 2020;371:m3862. – reference: 14. Wölfel R, Corman VM, Guggemos W, et al. Virological assessment of hospitalized patients with COVID-2019. Nature. 2020;581:465-469. – reference: 10. Schellenberg JJ, Ormond M, Keynan Y. Extraction-free RT-LAMP to detect SARS-CoV-2 is less sensitive but highly specific compared to standard RT-PCR in 101 samples. J Clin Virol. 2021;136:104764. – reference: 15. Yamada S, Fukushi S, Kinoshita H, et al. Assessment of SARS-CoV-2 infectivity of upper respiratory specimens from COVID-19 patients by virus isolation using VeroE6/TMPRSS2 cells. BMJ Open Respir Res. 2021;8:e000830. – ident: 13 doi: 10.1093/cid/ciaa1388 – ident: 7 doi: 10.1016/j.cca.2021.02.014 – ident: 5 doi: 10.1093/nar/28.12.e63 – ident: 11 doi: 10.1136/bmj.m3862 – ident: 10 doi: 10.1016/j.jcv.2021.104764 – ident: 9 doi: 10.1016/j.ijid.2020.08.029 – ident: 4 doi: 10.1128/JCM.00726-13 – ident: 15 doi: 10.1136/bmjresp-2020-000830 – ident: 14 doi: 10.1038/s41586-020-2196-x – ident: 2 doi: 10.20965/jdr.2021.p0084 – ident: 12 doi: 10.1016/j.jinf.2021.04.007 – ident: 1 doi: 10.1016/S0140-6736(20)30183-5 – ident: 8 – ident: 6 doi: 10.1016/j.jviromet.2006.11.031 – ident: 3 doi: 10.7883/yoken.JJID.2020.061 |
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| SubjectTerms | Antigens Chemiluminescence CLEIA Coronaviruses COVID-19 Enzyme immunoassay Gene amplification Immunoassay Polymerase chain reaction Reverse transcription Ribonucleic acid RNA RT-LAMP RT-PCR Saliva SARS-CoV-2 Severe acute respiratory syndrome coronavirus 2 Viral diseases |
| Title | Comparison of RT-PCR, RT-LAMP, and Antigen Quantification Assays for the Detection of SARS-CoV-2 |
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