Single-Domain Antibodies for the Detection of SARS-CoV-2 Nucleocapsid Protein.
Anal Chem
; 93(19): 7283-7291, 2021 05 18.
Article
in English
| MEDLINE | ID: covidwho-1217666
ABSTRACT
The goal of this work was to develop recombinantly expressed variable domains derived from camelid heavy-chain antibodies known as single-domain antibodies (sdAbs) directed against the SARS-CoV-2 nucleocapsid protein for incorporation into detection assays. To achieve this, a llama was immunized using a recombinant SARS-CoV-2 nucleocapsid protein and an immune phage-display library of variable domains was developed. The sdAbs selected from this library segregated into five distinct sequence families. Three of these families bind to unique epitopes with high affinity, low nM to sub-nM KD, as determined by surface plasmon resonance. To further enhance the utility of these sdAbs for the detection of nucleocapsid protein, homobivalent and heterobivalent genetic fusion constructs of the three high-affinity sdAbs were prepared. The effectiveness of the sdAbs for the detection of nucleocapsid protein was evaluated using MagPlex fluid array assays, a multiplexed immunoassay on color-coded magnetic microspheres. Using the optimal bivalent pair, one immobilized on the microsphere and the other serving as the biotinylated recognition reagent, a detection limit as low as 50 pg/mL of recombinant nucleocapsid and of killed virus down to 1.28 × 103 pfu/mL was achieved. The sdAbs described here represent immune reagents that can be tailored to be optimized for a number of detection platforms and may one day aid in the detection of SARS-CoV-2 to assist in controlling the current pandemic.
Full text:
Available
Collection:
International databases
Database:
MEDLINE
Main subject:
Camelids, New World
/
Single-Domain Antibodies
/
COVID-19
Type of study:
Experimental Studies
Limits:
Animals
/
Humans
Language:
English
Journal:
Anal Chem
Year:
2021
Document Type:
Article
Affiliation country:
Acs.analchem.1c00677
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