An ELISA for antibodies against infectious bronchitis virus using an S1 spike polypeptide.
Using the whole infectious bronchitis virus (IBV) for detecting the antibody against IBV by enzyme-linked immunosorbent assay (ELISA) is a routine work in poultry industry. To prepare virus is time consuming and tedious. Furthermore, the whole viral antigen detects all antibodies against the viral structural proteins, including spike (S), nucleocapsid, matrix, and other proteins. Among those, S protein is related to neutralization. Thus, to develop and express protein fragment from S gene and to use the protein as a coating antigen for antibody detection against IBV are the purposes of this experiment. A partial S gene fragment (n.t. 1143-1665) was cloned into pRSET vectors and transformed into competent Escherichia coli (E. coli) BL21 (DE3). A 27.5 kDa fusion protein (S-fg, containing S1-F and partial S2-G antigenic sites) was successfully expressed, affinity-purified and detected specifically with chicken anti-IBV serum by Western blot. The expressed S-fg protein was used as a coating antigen for developing an ELISA (S-fg ELISA) for serum antibody detection in anti-IBV antisera from different IBV serotypes and in field sera. The results show that the S-fg fusion protein is highly cross-reactive among different IBV serotypes, and the S-fg ELISA is found to be a convenient, economical, and efficient method for antibody detection against IBV.
Wang
C-H
CH
Department of Veterinary Medicine, National Taiwan University, P.O. Box 23-3, Taipei 106, Taiwan, ROC. chingho@ms.cc.ntu.edu.tw
Hong
C-C
CC
Seak
J C H
JC
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D015153:Blotting, Western; D002642:Chick Embryo; D002645:Chickens / Q000821:virology*; D003001:Cloning, Molecular; D018352:Coronavirus Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary*; D004926:Escherichia coli / Q000235:genetics; D001351:Infectious bronchitis virus / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D008562:Membrane Glycoproteins / Q000235:genetics / Q000276:immunology*; D011201:Poultry Diseases / Q000175:diagnosis* / Q000276:immunology / Q000821:virology; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D013047:Specific Pathogen-Free Organisms; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology*
D000914:Antibodies, Viral; D008562:Membrane Glycoproteins; D011994:Recombinant Proteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; C578557:spike glycoprotein, SARS-CoV; C578553:spike protein, mouse hepatitis virus