Development of blocking ELISA for detection of antibodies against avian influenza virus of the H7 subtype.
OBJECTIVE
The conventional method used for subtyping of antibodies against avian influenza viruses is hemagglutination inhibition (HI) test. However, the HI test is laborious and requires preparation of antigen from viable viruses that might be hazardous. The aim of this study was to develop a blocking enzyme-linked immunosorbent assay (B-ELISA) for detection of antibody of avian influenza of the H7 subtype. The B-ELISA is fast and avoids the need to culture whole viruses.


METHODS
The B-ELISA was based on the reaction between a monoclonal antibody and a recombinant hemagglutinin protein purified from Escherichia coli. The specificity of the B-ELISA was determined by testing H7-negative field sera and the sensitivity of the B-ELISA was determined by testing sera collected from experimentally immunized chickens.


RESULTS
The specificity of the B-ELISA was found to be 97.7% when compared with the HI test. The sensitivity was found to vary with the HI titer of sera. A sensitivity of 100% was achieved when test sera had HI titers >or=2(7). The sensitivity dropped to 33% and 20% when test sera had HI titers of 2(6) and 2(5), respectively. Nearly all test sera with HI titers <or=2(4) were scored negative by the B-ELISA.


CONCLUSIONS
The B-ELISA might serve as a useful tool for detection of H7-specific antibodies, with the added advantage that the recombinant hemagglutinin antigen could be produced in E. coli in large quantities, without handling the whole virus.
Shien
Jui-Hung
JH
Department of Veterinary Medicine, National Chung Hsing University, Taichung, Taiwan.
Fu
Lin-Fan
LF
Wu
Jin-Ru
JR
Cheng
Min-Chu
MC
Shieh
Happy K
HK
Chang
Poa-Chun
PC
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000914:Antibodies, Viral / Q000097:blood*; D002645:Chickens / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D009980:Influenza A virus / Q000235:genetics / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D012680:Sensitivity and Specificity
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D011994:Recombinant Proteins; C109963:hemagglutinin, avian influenza A virus