Antigenic mapping of the recombinant Norwalk virus capsid protein using monoclonal antibodies.
Norwalk virus (NV) is the prototype strain of a group of noncultivatable caliciviruses that infect humans and cause outbreaks of epidemic acute nonbacterial gastroenteritis. The NV virion is composed of 180 copies of a single structural protein that, when expressed in insect cells infected with a recombinant baculovirus, assembles into empty recombinant Norwalk virus-like particles (rNV VLPs) which are morphologically and antigenically similar to native NV. We have begun to dissect the antigenic structure of the rNV particles using monoclonal antibodies made to the rNV VLPs. Ten MAbs made to rNV particles were characterized for their reactivity as detector antibodies by ELISA, as capture antibodies in an ELISA to detect NV in stools, by Western blot, and by immunoprecipitation. Seven of the MAbs recognize discontinuous epitopes, requiring the rNV capsid protein to remain at least partially folded, while the other three recognize continuous epitopes. Eight of the MAbs map to the C-terminal half of the capsid protein as they react by Western blot and by immunoprecipitation with a 32K trypsin cleavage product of the full-length 58K capsid protein, suggesting that the C-terminal half of the capsid protein may contain the immunodominant epitopes. The three MAbs that recognize continuous epitopes map to the extreme C terminus of the capsid protein, between amino acids 457 and 530, in a region that is relatively conserved among different human calicivirus capsid proteins. These MAbs which were assigned into three antigenic groups will be useful as tools to further dissect the structural and antigenic topography of the NV virion, and as unlimited reagents to detect NV in diagnostic assays.
Hardy
M E
ME
Baylor College of Medicine, Houston, Texas 77030, USA.
Tanaka
T N
TN
Kitamoto
N
N
White
L J
LJ
Ball
J M
JM
Jiang
X
X
Estes
M K
MK
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000914:Antibodies, Viral / Q000276:immunology; D000956:Antigens, Viral / Q000276:immunology*; D015153:Blotting, Western; D002213:Capsid / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D005243:Feces / Q000821:virology; D006801:Humans; D051379:Mice; D009663:Norwalk virus / Q000276:immunology* / Q000648:ultrastructure; D011233:Precipitin Tests
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral