关键词: 间接免疫荧光组化; 禽波氏杆菌; 定位检测; 组织器官

Abstract: In order to study the colonization pattern,localization and quantity of Bordetella avium (B.avium) in the tissues and organs of chicks infected with B.avium,a method of indirect fluorescent immunohistochemistry suitable for B.avium detection was established and applied to detect tissues of artificially infected chicks in this study.Firstly the fixation agent for fixing tissues and organs of chicks was screened.Then the fixed tissues were sliced to sections and good pasting agent was detected.Rabbit anti-B.avium outer membrane protein IgG was used as the first antibody.After repeatedly optimizing the experiment conditions,10% neutral formalin was used as a fixation agent to fix tissues for 24 h and 0.1% poly-L-lysine as a section pasting agent.Optimal working conditions were as follows:2% skimmed milk powder closing section at 37℃ for 30 min;1:100 dilution of rabbit anti-B.avium outer membrane protein IgG incubating at 4℃ overnight;Fluorescein marked goat anti-rabbit secondary antibody was diluted 1:20,incubating at 37℃ for 30 min.This method had good reproducibility and specificity in B.avium detection.Detection results of chick tissues infected with B.avium showed that this method could specifically localize B.avium in the tissues and organs of chicks.Specific fluorescence signals could be found in tracheas lung,liver,heart,spleen and kidney.It had coincidence rate of 99.27% with bacterial isolation and culture.The results showed that the established indirect fluorescent immunohistochemistry could be used as an effective method in research of B.avium pathogenic mechanism.

Key words: indirect fluorescent immunohistochemistry; Bordetella avium; localization detection; tissues and organs