[Expression of recombinant VP1 protein of enterovirus 71 and development of serological assay for detection of EV71 infection].
OBJECTIVE
To obtain a recombinant purified Enterovirus 71 VPI protein and establishment of an early, rapid and accurate serological ELISA (enzyme-linked immunosorbent assay) for detection of EV71 infection.


METHODS
VP1 gene was amplified by PCR and clonel into pET-21b (+) vector, the positive recombinant plasmid were transformed into E. coli BI21(DE3), and was induced with IPTG, the recombinant protein by SDS-PAGE and Western Blot assays. Finally, the recombinant purified VP1 protein was used as a coated antigen for detection of serum anti-IgM and IgG against EV71 by ELISA.


RESULTS
The purified VP1 was obtained, and it can be recognized by sera of patients with EV71 infection associated with hand-foot-mouth disease. The A values of anti-EV71 IgM and IgG were significantly elevated as compared to healthy objects and HFMD patients without EV71 infection (P < 0.05). The sensitivity and specificity of IgM to EV71 were 73% and 77% compared with the RT-PCR results, respectively;and those of IgG being 82% and 83%, respectively.


CONCLUSIONS
The recombinant protein VP1 was produced and purified, and it was proved to have a good antigenicity and could be used to develop a serological diagnosis kit for EV71 infection in the future.
Zhou
Bo-ping
BP
The Third People's Hospital of Shenzhen, 518020, China. zhoubp@hotmail.com
Liu
Wei-long
WL
Xie
Jing-jing
JJ
Chen
Xin-chun
XC
Xu
Liu-mei
LM
Tan
Yan
Y
Liu
Ying-xia
YX
Yang
Gui-lin
GL
D000888:Antibodies, Anti-Idiotypic / Q000096:biosynthesis / Q000097:blood; D000914:Antibodies, Viral / Q000096:biosynthesis* / Q000276:immunology; D015153:Blotting, Western; D036022:Capsid Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D019411:Clinical Laboratory Techniques; D003001:Cloning, Molecular / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D004770:Enterovirus / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D015870:Gene Expression; D006232:Hand, Foot and Mouth Disease / Q000821:virology; D006801:Humans; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity
D000888:Antibodies, Anti-Idiotypic; D000914:Antibodies, Viral; D036022:Capsid Proteins; C442501:VP1 protein, Foot-and-mouth disease virus; C044203:anti-IgM