摘要
目的研制间日疟原虫乳酸脱氢酶(PvLDH)单克隆抗体,并对抗体特性进行了初步鉴定。方法用纯化的重组PvLDH蛋白免疫BALB/c小鼠,取其脾细胞与小鼠骨髓瘤细胞SP2/0进行融合,间接ELISA法筛选阳性杂交瘤细胞,测定其免疫球蛋白亚类及效价,结合ELISA、Western blot试验分析其特异性。结果共获得5株稳定分泌抗PvLDH重组蛋白单克隆抗体的杂交瘤细胞,分别为6D1、6D2、5A10、5C7、4A8。5株单抗培养上清的ELISA效价为1∶512～1∶1024,腹水效价为1∶12800～1∶25600,其中5C7、6D1经Western-blot鉴定能与天然的恶性疟原虫和间日疟原虫的LDH发生特异性反应。结论本研究方法成功制备并获得了针对PvLDH的单克隆抗体,为进一步研究疟疾诊断试剂奠定了基础。
Objective To prepare and characterize the monoclonal antibodies （McAbs） against lactate dehydrogen- ase of Plasmodium vivax （PvLDH）. Methods BALB/c mice were immunized with purified recombinant PvLDH. McAbs against PvLDH were prepared according to the protocol of hybridoma technique. Positive hybridoma cell lines were screened by ELISA. The McAbs were characterized by ELISA and Western blot analysis. Results Five hybridoma cell lines （6D1, 6D2, 5Al（）, 5C7 and 4F8） secreting MeAbs against recombinant PvLDH were obtained. All the 5 MeAbs were IgG1. The titers of these McAbs in supematant and ascites were 1：512 - 1 ： 1024 and 1 ： 12800 - 1：25600, respectively. Western blot a- nalysis revealed that two of the McAbs （5C7 and 6D1 ） strongly and specifically reacted with the native LDH in Plasmodium vivax and Plsmodiumfalciparum. Conclusion Two hybridoma cell lines secreting high titer of McAbs against PvLDH with high specificity have established. It provides a potential value for further research on the diagnosis of malaria.
出处
《广东医学》
CAS
CSCD
北大核心
2010年第8期942-944,共3页
Guangdong Medical Journal
基金
国家科技重大专项项目(编号:2008ZX10004-007)
广州市科技计划项目(编号:2002Z3-E4012)
广东省医学科研基金资助项目(编号:A2006365)
关键词
间日疟原虫
乳酸脱氢酶
单克隆抗体
plasmodium vivax
lactate dehydrogenase
monoclonal antibodies
