靶向造影剂(Gd-DTPA-Granzyme B)的制备  

Preparation of a novel targeted MR contrast agent Gd-DTPA-Granzyme B monoclonal antibody

作　　者：谢易[1] 刘广义[1] 韩飞[1] 姜虹[1] 茅幼英[1] 王慧萍[1] 赵杰[1] 金娟[1] 陈江华[1] 

机构地区：[1]浙江大学医学院附属第一医院肾脏病中心国家中医药管理局三级实验室"肾脏病免疫实验室",浙江杭州310003

出　　处：《浙江大学学报（医学版）》2010年第3期290-295,共6页Journal of Zhejiang University(Medical Sciences)

基　　金：国家自然科学基金(项目编号30671990);国家自然科学青年基金(项目编号30801148);115国家科技支撑计划(项目编号2008BAI60B04)

摘　　要：目的:探讨以颗粒酶B为靶向的磁共振造影剂(gadolinium-diethylenetriaminepentaacetic-granzyme B monoclonal antibody,Gd-DTPA-GB mAb)的制备方法及反应条件。方法:采用颗粒酶B单抗制成颗粒酶B靶向造影剂Gd-DTPA-GB mAb;通过基质辅助激光解析电离飞行时间质谱(matrix assisted laser desorption ionisation time-of-flight mass spectrometry,MALDI-TOF-MS)计算其相关参数,采用MTT法检测其细胞毒性,以及免疫组化法判定其免疫活性。结果:MALDI-TOF-MS鉴定结果显示:颗粒酶B单抗的质荷比高峰主要出现在133986.41处,连接Gd-DTPA后高峰主要在139736.06的位置;根据质谱结果计算得每分子抗体可以连接20个左右的Gd3+;MTT法结果显示:Gd、DTPA、Gd-DTPA,以及Gd-DTPA-GB mAb组在浓度逐渐递增的情况下对于细胞活性没有明显影响,且4组间比较差异无统计学意义(P>0.05);免疫组化结果显示:靶向造影剂(1∶175)与阳性对照组(1∶150)比较,具有较高的免疫活性。结论:在体外,本实验室设定的条件能够合成磁共振靶向造影剂Gd-DTPA-GB mAb,而且该造影剂具有良好的免疫活性,无明显细胞毒性。Objective： To prepare a novel MRI targeted contrast agent Gd-DTPA-Granzyme B monoclonal antibody（mAb） and to test its reaction conditions.Methods： The Granzyme B mAb was coupled with DTPA,and then conjugated with Gd.The Gd-DTPA antibody was characterized using MALDI-TOF-MS.Cytotoxicity test was performed with MTT assay,and immune activation was examined with immunohistochemistry.Results： MALDI-TOF-MS demonstrated that the molecular weight shifted from granzyme B mAb（133986） to Gd-DTPA-GB mAb（139736）,which indicated the conjugation of the antibody with Gd-DTPA.The molar ratio of Gd per IgG molecule was about 20.MTT assay showed that Gd,DTPA,Gd-DTPA and Gd-DTPA-GB mAb groups did not make an impact on cell viability,and there were no significant differences among 4 groups（P〉0.05）.Immunohistochemistry results showed that compared with the positive control group the targeted contrast agent had a high immune activity.Conclusion： The novel contrast agent Gd-DTPA-Granzyme B mAb prepared in this study keeps a good immune activity and has no significant cytotoxicity.

关 键 词：磁共振成像 三胺五乙酸 钆DTPA 造影剂 钆 光谱法 质量 基质辅助激光解吸电离 

分 类 号：TQ421.7[化学工程]