抗家蚕浓核症病毒单克隆抗体的制备及其在诊断上的应用

PRODUCTION OF MONOCLONAL ANTIBODIES TO DENSOVIRUS OF SILKWORM ( BOMBYX MORI ) AND THEIR APPLICATION IN DIAGNOSIS

获得6株能稳定分泌抗家蚕浓核症病毒(BmDNV)单克隆抗体(McAb)的Ⅰ杂交瘤细胞株(Bm(?)1～6)。其腹水型McAb经琼脂糖双向免疫扩散法检测,效价最高达256;用间接ELISA法测定,效价最高达250000.6株杂交癌细胞分泌的McAb均能有效地阻断BmDNV对家蚕的感染,中和指数以BmD_4最高,达1096。 建立了家蚕浓核症的抗原斑点酶标免疫方法,本法特异、灵敏、快速、简便,不仅可用于诊断,且可用于流行病学研究。

Monoclonal antibodies ( McAbs ) to densovirus ( DNV ) of silkworm ( Bombyx mori ) were generated by PEG fusion of splenocytes from immunized Balb/c mice and FO myeloma cells. After extensive screening, 6 hybridoma cell lines ( BmD1-6 ) were selected.IgG8 were secreted by BmD1,2,5, and IgG24, by BmD3, 4,6. The highest titer of ascites induced by these hybridoma cell lines was around 256 wfth double immunodifusion test and around 250 000 with ELISA indirect method. All McAbs secreted by these hybridoma cell lines can effectively neutralize the infectivity of BmDNV to silkworm. The highest neutralization index was around 1 096.McAbs Dot-ELISA direct, indirect, and sandwich methods were developed for the detection of BmDNV antigen from silkworm. The sensitivity of Dot-ELISA indirect method was 1 024 times that of double immunodifusion test, and 256 times that of countercurrect immunoelectrophoresis. The results demonstrated that these methods were specific, sensitive, fast and simple and may be used for the diagnosis and epidemiological studies of BmDNV