抗囊虫单抗的制备及其应用研究  

STUDY ON PREPARATION AND APPLICATION OF MONOCLONAL ANTIBODIES AGAINST CYSTICERCUS CELLULOSAE

作　　者：殷竹君[1] 张耀娟[1] 章子豪[1] 刘静[1] 史志明[1] 侯敏[1] 

机构地区：[1]南京医科大学寄生虫学教研室,江苏南京210029

出　　处：《中国寄生虫病防治杂志》2003年第2期77-80,共4页Chinese Journal of Parasitic Disease Control

基　　金：江苏省科委社会发展基金资助项目 (No.BS970 37)

摘　　要：目的　制备分泌特异性抗猪囊虫单抗的杂交瘤细胞株 ,建立以特异性单抗检测囊虫病患者循环抗原 (CAg)的特异敏感的 EL ISA方法。　方法　制备水溶性抗原 (Ag- w)和尿素溶性抗原 (Ag- u) ,将实验用 BAL B/ c小鼠分为 Ag- u和 Ag- w组 ,分别以 Ag- u和 Ag- w免疫 ,取两组免疫鼠脾细胞与 SP2 / 0骨髓瘤细胞融合 ,EL ISA法进行筛选 ,所获特异性阳性克隆加以扩大培养 ,并分别与包虫、血吸虫、华支睾吸虫、肺吸虫、姜片虫和丝虫抗原进行交叉反应性测定。　结果获得稳定分泌特异性抗囊虫单抗的杂交瘤细胞 5株 ,其中 1株 N1B10来自于 Ag- w组 ,另 4株 U2 A6、U2 B3、UB5 H6和UE10 H5来自于 Ag- u组。对 N1B10、U2 A6和 U2 B3初步鉴定结果表明 ,其抗体类型分别为 Ig G2 b、κ,Ig G1、κ,Ig G2 b、κ;腹水的效价分别为 1.0 2 4× 10 6、1.6× 10 4和 3.2× 10 4 ;免疫组化显示 N1B10定位于囊虫囊壁的基底膜 ,U2 A6和 U2 B3定位于囊虫的头节。杂交瘤细胞经连续培养 ,液氮反复冻存复苏后仍能稳定分泌单抗。取 3株单抗和兔抗囊虫血清建立了检测囊虫病患者 CAg的 EL ISA方法 ,测定 4 8份临床确诊囊虫病病人血清 ,阳性率达 95 .8%。　结论　成功制备了抗囊虫单克隆抗体 ,所建立的 EL ISA方法敏感性高 ,特异性强 ,可用于?Objective To prepare specific monoclonal antibody(McAb) against Cysticercus cellulosae and establish the ELISA method to detect CAg in sera of patients with cysticercosis. Methods To establish hybridoma cell lines secreting McAbs against C. cellulosae , water soluble and urea soluble antigens (Ag w and Ag u) from cysticercus were prepared. Immunization was carried out on two groups of BALB/c mice (group Ag w and group Ag u). After the third immunization, the spleen cells of the immunized mice were fused with SP2/0 myeloma cells. ELISA was applied to screen the McAbs that the hybridoma cells secreted. The positive cells were cloned and expanded immediately for the detection of cross reaction. Results One hybriodoma cell line secreting McAb named N1B10 from group Ag w and four hybrioloma cell lines secreting McAbs named U2A6, U2B3, UB5H6, UE10H5 from group Ag u were obtained. None of the five cross reacted with antigens of hydatid, schistosoma, clonochis sinensis, paragonimus, fasciolopsis, and filaria. N1B10, U2A6, U2B3 were identified as follows:The ELISA titers of N1B10, U2A6, U2B3 in the ascites were 1.024×10 6, 1.6×10 4 and 3.2×10 4 respectively . Ig class and subclass were IgG2b, k(N1B10), IgG1, k(U2A6), IgG2b, k(U2B3) separately. Immunohistochemical analysis indicated that N1B10 was located on the ground membrane of the cyst wall and U2A6, U2B3 were located on the scolex. The ELISA method established with McAbs and sera of the immunized rabbits was applied to detect CAg in sera from 48 patients with cysticercosis, the sensitivity was as high as 95.8%(46/48). Conclusion The specific McAbs against C. cellulosae were successfully prepared and the established ELISA method could be used to detect CAg in serum of patients with cysticercosis.

关 键 词：猪囊虫 尿素溶性抗原 水溶性抗原 单克隆抗体 ELISA 

分 类 号：R383.34[医药卫生—医学寄生虫学]