新城疫单克隆抗体的制备与特异性分析

Preparation and specificity analysis of monoclonal antibodies against Newcastle disease virus

以蔗糖反透析浓缩及ＳｅｐｈａｄｅｘＧ 200层析纯化Ｆ48Ｅ8和ＬａＳｏｔａ新城疫病毒(ＮＤＶ),取纯化的病毒液免疫接种ＢＡＬＢ ｃ小鼠,将小鼠脾细胞与ＮＳ 1骨髓瘤细胞融合,经次黄嘌呤、氨基喋呤和胸苷(ＨＡＴ)培养基选择,间接ＥＬＩＳＡ检测,有限稀释法克隆,筛选出了3株杂交瘤细胞,依次命名为1Ｄ4、1Ｄ5、16Ｇ12。3株杂交瘤细胞的染色体数为80～100,并能稳定传代,其产生的单克隆抗体重链属于ＩｇＧ1,轻链属于κ链;ＥＬＩＳＡ检测时仅与ＮＤＶ发生特异性反应,而与其他常见禽类病毒抗原不出现交叉反应,应用这些单克隆抗体对ＮＤＶ分离株进行了分群研究。

The chicken embryo allantoic fluid infected with NDV was concentrated by reverse dialysis with sucrose.The concentrated virus was purified by chromatography Sephadex G-200 column. Three strains of hybridoma cells secreting monoclonal antibody (McAb) were obtained by the fusion of NS-1 (myeloma) cell line with spleen cell from mice immunized with F_(48)E_(8) and LaSota viruses, and by cultivating the fusion cells in HAT medium, screening them with ELISA and cloning them with the limited dilution. The three strains of hybridoma cells were designated as 1D_4,1D_(5) and 16G_(12 ) respectively. The number of (chromosome) of the three strains of hybridoma cells was from 80 to 100.The McAbs were secreted from the three strains of hybridoma cells which were cultivated for two months. The H-chain of the three McAbs belonged to IgG_(1 )and L-chain belonged to κ type. They were specific for NDV and no cross reactions were (observed) with other avian virus antigens. The NDV isolates were divided into three groups with the three McAbs.