摘要
用硫酸铵分级盐析法纯化大菱鲆血清免疫球蛋白IgM,所得产物用Sepharose-4B和DEAE-Sepharose Fast Flow阴离子交换层析进一步纯化,以纯化的大菱鲆IgM免疫新西兰大白兔,获得兔抗大菱鲆IgM抗血清。SDS-PAGE电泳显示大菱鲆IgM重链为76 kD,轻链为27 kD;纯化的大菱鲆IgM重链与特异性抗血清具有较好的反应,而轻链与抗血清反应不明显;以制备的兔抗大菱鲆IgM抗血清为二抗建立了大菱鲆血清特异性抗体的间接ELISA检测方法,用该方法检测了鳗弧菌灭活疫苗免疫后大菱鲆产生特异性抗的变化规律,大菱鲆在免疫后第1周就产生了特异性抗体,在3周时达到峰值,该特异性抗体可维持13周以上。
An immunoglobulin M （IgM） was purified from serum of turbot with ammonium sulfate precipitation, followed with Sepharose-4B column chromatography and DEAE-Sepharose Fast Flow column chro-matogsaphy. SOS-PAGE eleetrophoresis showd that the purified lgM was composed of 76 kD heavy chains and 27 kD light chains. Antiserum against the purified IgM was prepared by repeatedly immunized New Zealand rabbits with the purified protein. Western blot analysis showed that the heavy chain of the purified IgM reacted well with the rabbit antiserum while the light chain hardly reacted with it. An indirect ELISA method was established using the specific antiserum as second antibody. Turbot was immunized with formalin-killed Vibrio anguillarum cells via intraperitoneal injection. The specific antibody was determined with the indirect ELISA. Specific antibody could be detected in a week after immunization. The titer of the antibody reached to a peak in three weeks, and maintained a high level till thirteen weeks.
出处
《中国海洋大学学报（自然科学版）》
CAS
CSCD
北大核心
2008年第6期932-936,共5页
Periodical of Ocean University of China
基金
国家自然科学基金项目(30371108)
国家高技术研究发展计划项目(2003AA622070,2007AA09Z416)资助
关键词
大菱鲆
IGM
纯化
抗血清
ELISA
turbot （Scophthalmus maximus ）
immunoglobulin M （IgM）
purification
anti-serum
ELISA
