关键词: 禽流感, NS1蛋白, 胶体金试纸条

Abstract:

【Objective】 A rapid assay based on colloidal gold immunochromato graphic method was developed to identify AIV-infected chickens from those immunized with invactivated vaccine, which was charactered with rapid and simple handling on spot by even unskilled feeder. 【Method】 The NS1 gene of H9N2 subtype influenza A virus was cloned into vector pGEX-KG to construct the recombinant plasmid KG-NS1, which was then transformed into E. coli BL21-DE3 competent cells and induced by IPTG. After analysed by western-blot, the purified product was used as diagnostic antigen in development of a colloidal gold immunochromatographic assay (GICA) strip for the NS1 antibody detection. The specificity and sensitivity of the strip were evaluated, and the sera from experimental and clinical samples were tested. 【Result】 The fusion protein GST-NS1 (r-NS1) was about 52 kD and showed immunologic competence. Using the purified r-NS1, a GICA strip was developed for the NS1 antibody detection with high specificity and acceptable sensitivity, whose results were decided through the observation of the test line in red or not (red indicated positive result) by naked eye within 25 min. Tested by the strip, only the sera of r-NS1 protein and AIV-infected read positive, but the sera of other antigen read negative. Using the GICA strip, the NS1 antibody of the experimental AIV-infected chickens was identified. The NS1 antibodies were detected as early as the third day post- infection and maintained for around two weeks. It was about 80% experimental sample and 9.1% clinical samples detected positive with the strip. 【Conclusion】 GICA is a visual, specific, rapid and simple detection method and it is feasible to apply to distinguish between vaccinated and virus-infected poultry in field screening. It is worth to be popularized.

Key words: avian influenza, NS1, colloidal gold strip