Development of a highly specific and sensitive VHH-based sandwich immunoassay for the detection of the SARS-CoV-2 nucleoprotein.
J Biol Chem
; 298(1): 101290, 2022 01.
Article
in English
| MEDLINE | ID: covidwho-1472024
ABSTRACT
The current COVID-19 pandemic illustrates the importance of obtaining reliable methods for the rapid detection of SARS-CoV-2. A highly specific and sensitive diagnostic test able to differentiate the SARS-CoV-2 virus from common human coronaviruses is therefore needed. Coronavirus nucleoprotein (N) localizes to the cytoplasm and the nucleolus and is required for viral RNA synthesis. N is the most abundant coronavirus protein, so it is of utmost importance to develop specific antibodies for its detection. In this study, we developed a sandwich immunoassay to recognize the SARS-CoV-2 N protein. We immunized one alpaca with recombinant SARS-CoV-2 N and constructed a large single variable domain on heavy chain (VHH) antibody library. After phage display selection, seven VHHs recognizing the full N protein were identified by ELISA. These VHHs did not recognize the nucleoproteins of the four common human coronaviruses. Hydrogen Deuterium eXchange-Mass Spectrometry (HDX-MS) analysis also showed that these VHHs mainly targeted conformational epitopes in either the C-terminal or the N-terminal domains. All VHHs were able to recognize SARS-CoV-2 in infected cells or on infected hamster tissues. Moreover, the VHHs could detect the SARS variants B.1.17/alpha, B.1.351/beta, and P1/gamma. We propose that this sandwich immunoassay could be applied to specifically detect the SARS-CoV-2 N in human nasal swabs.
Keywords
Full text:
Available
Collection:
International databases
Database:
MEDLINE
Main subject:
Enzyme-Linked Immunosorbent Assay
/
Nucleocapsid Proteins
/
Single-Domain Antibodies
/
SARS-CoV-2
Type of study:
Diagnostic study
Topics:
Variants
Limits:
Animals
/
Humans
Language:
English
Journal:
J Biol Chem
Year:
2022
Document Type:
Article
Affiliation country:
J.jbc.2021.101290
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