摘要
鸽卵黄经硫酸铵盐析、酒精沉淀及凝胶 ( Sephadex G- 1 0 0 )柱层析后 ,得到较纯的鸽Ig G。以其免疫 BAL B/C小鼠 ,取其脾细胞与骨髓瘤细胞 ( SP2 /0 )进行细胞融合 ,经过酶联免疫吸附试验 ( ELISA)筛选 ,得到两株能稳定分泌抗鸽 Ig G的单克隆抗体 (以下称单抗 ) ,分别命名为 1 E5和 4C2。经亚类鉴定 ,该单抗均为 Ig G1亚类。在鸽 Ig G包被的 96孔板上用间接 ELISA方法测定单抗腹水的 ELISA效价 ,结果表明 ,单抗 1 E5和 4C2的 ELISA效价分别为 1∶ 1 60 0和 1∶ 640 0。免疫印迹试验结果表明 ,两株单抗均识别 Ig G重链 (分子量约为 66KD)。将单抗分别与鸡、鸭、鹅、鹌鹑和鸽等禽类血清包被的酶标板作间接 ELISA,结果表明单抗只与鸽血清的蛋白发生特异性反应 ,而不与其它禽类血清的蛋白反应。这说明抗鸽 Ig
Pigeon IgG,which was purified from pigeon egg yolk by water dilution,(NH 4) 2SO 4 precipitation,alcohol precipitation and column chromatography with sephadex G\|100,was used as antigen to immune BALB/C mice. By fusion of SP2/0 myeloma cells and spleen cells of these mice,two monoclonal antibodies against pigeon IgG in indirect ELISA were screened and designated as IE5 and 4C2.Both of these two belonged to IgG 1 subclass, and reacted with an approximately 66KD protein band in Western blotting,indicating that they recognized the heavy chains of pigeon IgG.In indirect ELISA, the titers of IE5 and 4C2 ascitic fluids were 1∶1600 and 1∶6400 respectively, and they reacted only with the pigeon serum and pigeon IgG but not the sera of other avian species, such as chicken , duck ,quail , goose.This is the first report concerning the preparation of the monoclonal antibody against pigeon IgG in China. They could be used as a common reagent in assay for detecting antibody levels to different pathogen in pigeon sera. It has potential to be used to develop a diagnostic kit.
出处
《中国兽药杂志》
北大核心
2000年第2期9-13,共5页
Chinese Journal of Veterinary Drug
