摘要
为开展弓形虫免疫诊断及抗弓形虫感染的相关研究,利用可溶性表达的弓形虫表面抗原SAG1截短型重组蛋白(trSAG1)研制单克隆抗体,筛选出能够稳定分泌高滴度抗体的杂交瘤细胞株,所分泌的单抗不仅适用于在虫体定位该蛋白,而且具有一定的小鼠体内保护作用。共筛选出4株(6H1、6G12、7B4和7A12)特异性好的单抗分泌株,经亚型测定,4株单抗重链均为IgG1,轻链均为κ链。抗体经纯化后测定效价为1∶8 000～1∶25 600;Western blot结果显示4株单抗能够同时识别重组表达及天然弓形虫SAG1抗原。筛选得到的2株抗SAG1单抗(6H1和7B4)可用于间接免疫荧光检测SAG1蛋白在虫体中的表达定位和分布。小鼠被动免疫保护试验表明,6H1和7B4两株单抗分别或联合使用可稍微延长被弓形虫感染的小鼠的存活时间。本试验研制的弓形虫SAG1单克隆抗体对将来研究弓形虫蛋白定位、弓形虫感染的诊断及防治具有一定的应用价值。
To develop the detection methods for Toxoplasma gondii, monoclonal antibodies （McAbs） to T. gondii major surface antigen SAG1 were generated, and screened and identified. Four strains of hybridoma cells including 6H1, 6G12, 71M and 7A12 which had good specificity against T. gondii were obtained. The subtypes of all the four McAbs were determined. The heavy chains were IgG1 and the light chains were K-chain. Four McAbs were purified and the titers were from 1 ： 8 000 to 1 ： 25 600. Western blot analysis demonstrated that all the four McAbs reacted strongly and specifically with recombinant and tachyzoite native SAG1 antigen of T. gondii, but not with pET Trx pro- tein. McAbs 6H1 and 7B4 were applicable for the immunofluorescent observation of SAG1 in tachyzoite. The immune protection tests also in- dicated that both 6H1 and 7B4 could prolong the survival time of the challenged mice. Our results indicated that the two McAbs will be useful in further studies for SAG1 location in T. gondiii, and diagnosis and treatment of toxoplasmosis.
出处
《畜牧与兽医》
北大核心
2013年第5期9-13,共5页
Animal Husbandry & Veterinary Medicine
基金
中央级公益性科研院所基本科研业务费专项资金(2013JB10)
公益性行业(农业)科研专项经费(201303045)
关键词
弓形虫
SAG1
单克隆抗体
免疫荧光检测
免疫保护
Toxoplasma gondii
major surface antigen SAG1
monoclonal antibody
immunofluorescent observation
protective immunity
