TY - JOUR
T1 - Proposal of De Novo Antigen Test for COVID-19
T2 - Ultrasensitive Detection of Spike Proteins of SARS-CoV-2
AU - Kyosei, Yuta
AU - Namba, Mayuri
AU - Yamura, Sou
AU - Takeuchi, Rikiya
AU - Aoki, Noriko
AU - Nakaishi, Kazunari
AU - Watabe, Satoshi
AU - Ito, Etsuro
N1 - Funding Information:
Funding: This research was funded by the A-STEP Program from JST (grant number AS3015096U), the Matching Planner Program from JST (grant number VP29117939087), the Development of Systems and Technology for Advanced Measurement and Analysis from JST, the JSPS KAKENHI (grant number 20H04556), the Waseda University grants for Specific Research Projects (grant numbers 2017A-015 and 2019C-123), and the Precise Measurement Technology Promotion Foundation.
Publisher Copyright:
© 2020 by the authors.
PY - 2020/8
Y1 - 2020/8
N2 - Polymerase chain reaction (PCR)-based antigen tests are technically difficult, time-consuming, and expensive, and may produce false negative results requiring follow-up confirmation with computed tomography. The global coronavirus disease 2019 (COVID-19) pandemic has increased the demand for accurate, easy-to-use, rapid, and cost-effective antigen tests for clinical application. We propose a de novo antigen test for diagnosing COVID-19 using the combination of sandwich enzyme-linked immunosorbent assay and thio-nicotinamide adenine dinucleotide (thio-NAD) cycling. Our test takes advantage of the spike proteins specific to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus. The limit of detection of our test was 2.3 × 10−18 moles/assay. If the virus has ~25 spike proteins on its surface, our method should detect on the order of 10−20 moles of virus/assay, corresponding to ~104 copies of the virus RNA/assay. The detection sensitivity approaches that of PCR-based assays because the average virus RNA load used for PCR-based assays is ~105 copies per oro- or naso-pharyngeal swab specimen. To our knowledge, this is the first ultrasensitive antigen test for SARS-CoV-2 spike proteins that can be performed with an easy-to-use microplate reader. Sufficient sensitivity can be achieved within 10 min of thio-NAD cycling. Our antigen test allows for rapid, cost-effective, specific, ultrasensitive, and simultaneous multiple measurements of SARS-CoV-2, and has broad application for the diagnosis for COVID-19.
AB - Polymerase chain reaction (PCR)-based antigen tests are technically difficult, time-consuming, and expensive, and may produce false negative results requiring follow-up confirmation with computed tomography. The global coronavirus disease 2019 (COVID-19) pandemic has increased the demand for accurate, easy-to-use, rapid, and cost-effective antigen tests for clinical application. We propose a de novo antigen test for diagnosing COVID-19 using the combination of sandwich enzyme-linked immunosorbent assay and thio-nicotinamide adenine dinucleotide (thio-NAD) cycling. Our test takes advantage of the spike proteins specific to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus. The limit of detection of our test was 2.3 × 10−18 moles/assay. If the virus has ~25 spike proteins on its surface, our method should detect on the order of 10−20 moles of virus/assay, corresponding to ~104 copies of the virus RNA/assay. The detection sensitivity approaches that of PCR-based assays because the average virus RNA load used for PCR-based assays is ~105 copies per oro- or naso-pharyngeal swab specimen. To our knowledge, this is the first ultrasensitive antigen test for SARS-CoV-2 spike proteins that can be performed with an easy-to-use microplate reader. Sufficient sensitivity can be achieved within 10 min of thio-NAD cycling. Our antigen test allows for rapid, cost-effective, specific, ultrasensitive, and simultaneous multiple measurements of SARS-CoV-2, and has broad application for the diagnosis for COVID-19.
KW - COVID-19
KW - SARS-CoV-2
KW - Spike protein
KW - Thio-NAD cycling
KW - Ultrasensitive ELISA
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U2 - 10.3390/diagnostics10080594
DO - 10.3390/diagnostics10080594
M3 - Article
AN - SCOPUS:85090191595
SN - 2075-4418
VL - 10
JO - Diagnostics
JF - Diagnostics
IS - 8
M1 - 594
ER -