Study On Infectious Bronchitis Virus And Anti-Ibv Antibody In Chicken By Silver-Enhanced Colloidal Gold Assay

The purified goat anti-rat IgG or rabbit anti-chicken IgG was labeled with colloidal gold particals of 10nm in diameter. A Silver-Enhanced Colloidal Gold Assay was developed for detection of IBV and anti-IBV antibody in chickenWhen IBV was detected , nitrocellulose membrane (NCM) with IBV to be detected was blocked and reacted in solution of anti-IBV McAb diluted with 1:40 for 10 minutes. After being washed, NCM was immersed in the solution of goat anti-rat IgG labeled with collidal gold diluted with 1:4 for one hour. Then NCM with sample was stained 10 minutes by silver. The sample with clear and obvious black dot was judged as positive reaction for IBV, otherwise, negative result. The detection limit of purifiedIBV antigen and allantoic fluid of IBV were 0.703ng with each dot (0.3515ug/ml), 102.9 EID50 respectively. IBV titer in inoculated eggs was highest from 36 to 54 hours post inoculation (p. i.). IBV could be detected in 3 days p.i. in trachea of specific pathogen free (SPF) chicks by SECGA at least. When anti-IBV antibody in chicken was detected, first purified IBV was coated onto NCM with 0.4 u g each dot. After being blocked , the NCM was immersed in serum samples diluted with different titer (10x2x) for 10 minutes and then reacted in solution of rabbit anti-chicken IgG labeled with colloidal gold diluted with 1:4 for one hour .In the end NCM was stained 10 minutes by silver and the result was observed. IBV positive serum titer was more than 10 X 27> whereas ND, IBD, EDS.76 positive serum titer was less than 10 X21"2, which indicated crossreaction was no signification. The detection limit of chicken IgG was 0.88ng each dot. SECGA, Serum Neutralization in IOCS and ELISA were significantly corelative when anti-IBV antibody was detected using them respectively, when SECGA titer of serum was 1:160 , ELISA titer (P/N) was 2. So serum sample was positive as serum SECGA titer was more than 1:160 On the contrary the serum was negative . SECGA could be applied to diagnose IBV early andused for antibody surveillance of IB .It was group - specific, rapid, sensitive, simple. What's more it needed no particular instruments and the result was determined easily and so on .SECGA could be performed especially in clinical diagnosis and primary laboratory...