马铃薯X病毒云南分离物单克隆抗体的制备及其检测应用

Production and application of monoclonal antibodies against Potato virus X

用马铃薯X病毒(Potato virusX,PVX)免疫的BALB/C鼠脾细胞与SP2/0鼠骨髓瘤细胞融合,经筛选克隆,获得2株能稳定传代并分泌抗PVX单克隆抗体(MAb)的杂交瘤细胞2B12和4E7,并分别制备它们的单抗腹水.2株单克隆抗体腹水间接ELISA效价在10-6以上,2B12和4E7的抗体类型及亚类均为IgG1,轻链均为κ链.利用这些单克隆抗体建立了抗原包被的间接ELISA(ACP-ELISA)检测PVX的方法.病叶经1∶600倍稀释、提纯PVX病毒浓度为2 ng/mL(每孔的病毒绝对量为0.2 ng)时,该方法仍能检测到病毒.利用ACP-ELISA方法测定了田间样品,发现PVX在马铃薯上发病很普遍.

Two hybridoma cell lines, 2B12 and 4E7, secreting monoclonal antibodies （MAbs） against Potato virus X （PVX） were produced by fusing mouse myeloma cells （SP2/0） with spleen cells from BALB/C immunized by the PVX particles. The two MAbs could react specifically with PVX and the titres of their ascitic fluids were up to 10.6 in ELISA. Isotypes and subclasses of both 2B12 and 4E7 were found to be allied to IgG1. When used in antigen-coated plate （ACP） -EI.ISA for PVX detection, these two MAbs could successfully detect purified PVX at the amount as low as 0.2 ng or virus in plant sap diluted to a ratio of 1：600. ACP ELISA based on two MAbs was used to investigate the presence of PVX in field potato and results obtained indicate that PVX is widespread in field potatoes.