温度对牙鲆皮肤黏液抗体产生的影响

在9℃、15℃、21℃和26℃4种不同水温下,用淋巴囊肿病毒(LCDV)灭活疫苗腹腔注射牙鲆,应用间接酶联免疫吸附试验(ELISA)研究了其皮肤黏液中特异性抗体水平的变化,以分析温度对牙鲆皮肤黏液抗体产生的影响.ELISA结果表明,9℃和15℃水温下牙鲆黏液OD值分别在注射LCDV后第9周和第7周达到峰值(9℃:OD=0.179; 15℃:OD=0.233); 21℃水温下OD值上升最快,5周达到峰值(0.316); 26℃水温下OD值较21℃无显著差异,也于5周达到峰值(0.295).采用硫酸铵分步盐析等技术粗提不同温度下OD值最高时的牙鲆皮肤黏液中的免疫球蛋白(Ig),SDS-PAGE检测发现,各温度组黏液蛋白中均含有72 kD和26 kD蛋白条带.Western blotting结果显示,抗牙鲆血清Ig重链的单克隆抗体只与黏液蛋白中72 kD条带发生反应,确定为牙鲆皮肤黏液Ig重链.综上结果表明,牙鲆在最适生活温度(21℃)下,抗体应答强度最大.牙鲆粗提黏液蛋白中Ig的初步确定,为探索牙鲆黏液免疫机制提供了材料.

Effect of temperature on production of skin mucus antibody in flounder (Paralichthys olivaceus)

We evaluated the effects of water temperature on the kinetics of skin mucus antibody in the flounder sing an indirect enzyme-linked immunosorbent assay (ELISA). The fish were held at 9 and given an intraperitoneal injection of inactivated lymphocystis disease virus (LCDV). The optical density (OD) of mucus antibody peaked 9 and 7 weeks after treatment in the groups held at 9 during the 5th week. The OD was lower in fish held at 26) in the 5th week. We collected a sample of skin mucus from fish with the highest OD and extracted mucus Ig by ammonium sulfate precipitation. The mucus proteins from all four treatment groups yielded bands at 72 kD and 26 kD. Western blotting analysis revealed that the monoclonal antibody against serum Ig heavy chain could only recognize the 72 kD protein, which represents the heavy chain of skin mucus Ig. In conclusion, the most pronounced immune response was measured in fish that were maintained at their optimal temperature (21