Generation of human antibody fragments recognizing distinct epitopes of the nucleocapsid (N) SARS-CoV protein using a phage display approach.
BACKGROUND
Severe acute respiratory syndrome (SARS)-CoV is a newly emerging virus that causes SARS with high mortality rate in infected people. Successful control of the global SARS epidemic will require rapid and sensitive diagnostic tests to monitor its spread, as well as, the development of vaccines and new antiviral compounds including neutralizing antibodies that effectively prevent or treat this disease.


METHODS
The human synthetic single-chain fragment variable (scFv) ETH-2 phage antibody library was used for the isolation of scFvs against the nucleocapsid (N) protein of SARS-CoV using a bio panning-based strategy. The selected scFvs were characterized under genetics-molecular aspects and for SARS-CoV N protein detection in ELISA, western blotting and immunocytochemistry.


RESULTS
Human scFv antibodies to N protein of SARS-CoV can be easily isolated by selecting the ETH-2 phage library on immunotubes coated with antigen. These in vitro selected human scFvs specifically recognize in ELISA and western blotting studies distinct epitopes in N protein domains and detect in immunohistochemistry investigations SARS-CoV particles in infected Vero cells.


CONCLUSIONS
The human scFv antibodies isolated and described in this study represent useful reagents for rapid detection of N SARS-CoV protein and SARS virus particles in infected target cells.
Flego
Michela
M
Department of Drug Research and Evaluation, Istituto Superiore di Sanità, Rome, Italy. flego@iss.it
Di Bonito
Paola
P
Ascione
Alessandro
A
Zamboni
Silvia
S
Carattoli
Alessandra
A
Grasso
Felicia
F
Cassone
Antonio
A
Cianfriglia
Maurizio
M
D000914:Antibodies, Viral / Q000276:immunology*; D000086462:Coronavirus Nucleocapsid Proteins; D000939:Epitopes / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007128:Immunoglobulin Fragments / Q000276:immunology*; D019590:Nucleocapsid Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D019151:Peptide Library; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics / Q000276:immunology*
D000914:Antibodies, Viral; D000086462:Coronavirus Nucleocapsid Proteins; D000939:Epitopes; D007128:Immunoglobulin Fragments; C000706247:N protein, SARS-CoV; D019590:Nucleocapsid Proteins; D019151:Peptide Library; C055966:immunoglobulin Fv