Development and characterization of monoclonal antibodies against avian reovirus sigma C protein and their application in detection of avian reovirus isolates.
Avian reovirus (ARV) is a non-enveloped virus with a segmented double-stranded RNA genome surrounded by a double icosahedral capsid shell. ARVs are associated with viral arthritis, immunosuppression, and enteric diseases in poultry. The sigma C protein was involved in induction of apoptosis and neutralization antibody. In the present study, sigma C-His protein was expressed in Sf9 insect cells and purified by immobilized metal affinity chromatography. Eight monoclonal antibodies (mAbs) against sigma C-His and three mAbs against His were screened from hybridoma cells produced by fusion of splenocytes from immunized mice with NS1 myeloma cells. Among the eight mAbs against sigma C protein, all belonged to the IgG isotype except three for IgM. It was discovered that all anti-His mAbs were mixtures of IgG and IgM isotypes. mAbs reacted with sigma C-His protein in a conformation-independent manner based on dot blot and western blotting assays. The competitive binding assay indicated that all mAbs recognized the same epitope on sigma C protein that was conserved in different isolates. Compared with the commercial anti-ARV S1133 polyclonal antibody, mAb (D15) had universal reactivity to all serotypes or genotypes of ARVs tested. This monoclonal antibody may therefore be useful for the development of an antigen-capture enzyme-linked immunosorbent assay for rapid detection of field isolates.
Hsu
Chien J
CJ
Department of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung, Taiwan.
Wang
Chi Y
CY
Lee
Long H
LH
Shih
Wen L
WL
Chang
Chi I
CI
Cheng
Hsueh L
HL
Chulu
Julius L C
JL
Ji
Wen T
WT
Liu
Hung J
HJ
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000915:Antibody Affinity; D036022:Capsid Proteins / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary; D018604:Epitope Mapping; D051379:Mice; D008807:Mice, Inbred BALB C; D030101:Orthoreovirus, Avian / Q000276:immunology* / Q000302:isolation & purification*; D014764:Viral Proteins / Q000276:immunology; D054334:Viral Regulatory and Accessory Proteins
D000911:Antibodies, Monoclonal; D036022:Capsid Proteins; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C031728:sigma 1 protein, reovirus; C031657:sigma NS protein, Reovirus