[Purification and reactivity of foot-and-mouth disease virus non-structural protein 3A, 3B and 2C expressed in E. coli].
OBJECTIVE
To purify and to detect reactivity of non-structural proteins 3A, 3B and 2C expressed in the Escherichia coli.


METHODS
FMDV NSP 3A, 3B and 2C containing the major B-cell antigenic sites were expressed in E. coli. We got renatured 2C protein by lysing of isolated inclusion body using high concentration of urea, and then diluted in a buffer system containing oxidized/reduced glutathione. Purified 3A, 3B and 2C were obtained by Ni-NTA His Bind Resin affinity chromatography. The reactivity of three NSPs with sera of different origin was measured using an indirect ELISA and Western-blot. The reactivity of three proteins was compared with 3ABC and 3D by detecting sera of clinically healthy sheep that were collected from epidemic region of Asia I FMD.


RESULTS
Proteins 3A and 3B were solubly expressed in bacteria, and 2C was expressed to form inclusion body. All three products could react specifically with sera from FMDV infected animal by western-blot and ELISA. The high coincident rates were observed between 3A, 3B, 2C and 3ABC.


CONCLUSIONS
The results would provide useful materials for establishment of immunoelectro-transfer blot (EITB) diagnostic method, which could be used for differentiation of the FMDV infected animals from the vaccinated animals.
Fu
Yuanfang
Y
State Key Laboratory of Veterinary Etiological Biology, National Foot-and-Mouth Disease Reference Laboratory, Lanzhou Veterinary Research Institute of Chinese Academy of Agriculture Science, Lanzhou, China. wst258@163.com
Lu
Zengjun
Z
Cao
Yimei
Y
Guo
Jianhong
J
Zhang
Xiaoli
X
Tian
Meina
M
Liu
Zaixin
Z
Cai
Xuepeng
X
D000818:Animals; D000918:Antibody Specificity; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics*; D029544:Foot-and-Mouth Disease Virus; D015870:Gene Expression; D007106:Immune Sera / Q000032:analysis / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D012995:Solubility; D017361:Viral Nonstructural Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*
D007106:Immune Sera; D011993:Recombinant Fusion Proteins; D017361:Viral Nonstructural Proteins