Identification of enteroviruses by using monoclonal antibodies against a putative common epitope.
A common epitope region of enteroviruses was identified by sequence-independent single-primer amplification (SISPA), followed by immunoscreening of 11 cDNA libraries from two Korean enterovirus isolates (echoviruses 7 and 30) and a coxsackievirus B3 (ATCC-VR 30). The putative common epitope region was localized in the N terminus of VP1 when the displayed recombinant proteins from the phages were chased by the convalescent-phase sera. The genomic region encoding the common epitope region was amplified and then expressed by using the vector pGEX-5X-1. The antigenicity of the expressed recombinant protein was identified by Western blotting with guinea pig antisera for six different serotypes of enteroviruses. After successive immunization of mice with the recombinant common epitope protein, splenocytes were extracted and hybridized with P3X63-Ag8-653 cells. A total of 24 hybridomas that produced monoclonal antibodies (MAbs) against the putative common epitope of enteroviruses were selected. Four of these were immunoglobulin G1 isotypes with a kappa light chain. These MAbs recognized 15 Korean endemic serotypes and prototypes of enteroviruses in an indirect immunofluorescence assay. These results suggest that the expressed protein might be a useful antigen for producing group common antibodies and that the use of the MAbs against the putative common epitope of enteroviruses might be a valuable diagnostic tool for rapidly identifying a broad range of enteroviruses.
Shin
Soo-Youn
SY
Department of Life Science, College of Natural Science, Chung-Ang University, Seoul 156-756, Korea.
Kim
Ki-Soon
KS
Lee
Yoon-Sung
YS
Chung
Yoon-Seok
YS
Park
Kwi-Sung
KS
Cheon
Doo-Sung
DS
Na
Byoung-Kuk
BK
Kang
Yoonsung
Y
Cheong
Hyang-Min
HM
Moon
Youngjoon
Y
Choi
Jee-Hye
JH
Cho
Hang-Eui
HE
Min
Na-Young
NY
Son
Jin-Sook
JS
Park
Young-Hoon
YH
Jee
Youngmee
Y
Yoon
Jae-Deuk
JD
Song
Chul-Yong
CY
Lee
Kwang-Ho
KH
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000914:Antibodies, Viral / Q000276:immunology*; D000918:Antibody Specificity; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D004770:Enterovirus / Q000145:classification* / Q000276:immunology; D000939:Epitopes / Q000276:immunology*; D005260:Female; D006801:Humans; D006825:Hybridomas; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques; D014764:Viral Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D017931:DNA Primers; D018076:DNA, Complementary; D000939:Epitopes; D014764:Viral Proteins