小鼠主要组织相容性复合物移植抗原肽的提取与验证  

Collection and verification of murine allo-transplantation major histocompatibility complex peptides

作　　者：周景师[1] 李霄[1] 李伟民[1] 纪洪辰 

机构地区：[1]第四军医大学附属西京医院肝胆外科暨器官移植中心,陕西西安710032

出　　处：《细胞与分子免疫学杂志》2015年第8期1009-1012,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：陕西省科学技术研究发展计划项目(2011K12-01-06)

摘　　要：目的建立主要组织相容性复合物(MHC)移植抗原肽提取方法。方法以供体C57BL/6小鼠脾细胞为移植抗原免疫BALB/c受体小鼠,用p H3.3柠檬酸缓冲液洗脱受体小鼠脾细胞MHC结合抗原肽,用BCA蛋白测定法和高效液相色谱串联质谱法(HPLC-MS/MS)检测洗脱液中多肽成分,以小鼠异位心脏移植验证洗脱液内多肽的移植免疫抗原性。结果弱酸洗脱获得了微量多肽50μg/108脾细胞,其中含有混杂短多肽。与同系移植组相比,HPLC-MS/MS检测发现同种异体移植组洗脱液中出现差异多肽。同系MHC移植抗原肽预致敏受体心脏移植物中位存活时间为8 d,同种异体MHC移植抗原肽预致敏受体心脏移植物中位存活时间仅4 d。用同种异体来源的MHC移植抗原肽预致敏受体,使小鼠心脏移植物中位存活时间显著缩短。结论通过供体脾细胞免疫,弱酸洗脱受体脾细胞可获得MHC移植抗原肽,可用于进一步筛选移植抗原决定簇。Objective To establish a method of collecting transplantation major histocompatibility complex（ MHC）peptides. Methods Splenic cells of C57 BL /6 donor mice were injected into BALB/c recipient mice. The splenic cel s of the recipient mice were soaked in p H3. 3 citric acid buffer to wash off the MHC conjugated peptides. Peptide contents and components in the elutriant were detected by BCA protein assay and high performance liquid chromatography-tandem mass spectrometry（HPLC-MS/MS）. Antigenicity of the peptides was verified by murine heterotopic heart transplantation. Results Fifty micro-gram mixed peptides（ including short peptides） per 10^8 cells were collected by the elution method. HPLC-MS / MS revealed that there were differences in peptide components between allo-transplantation group and control group. Medial survival time of heart grafts of same strain MHC peptide pre-immunization group was 8 days,and that of allo-MHC peptide pre-immunization group was 4 days. Medial survival time of heart grafts was shortened significantly by pre-immunization with allo-MHC peptides. Conclusion Transplantation MHC peptides can be obtained by donor splenic cell immunization and elution from recipients＇ splenic cells with mild acid. They can be used for screening transplantation antigenic determinants.

关 键 词：移植免疫 抗原 抗原决定簇 T淋巴细胞 主要组织相容性复合物 

分 类 号：R392.11[医药卫生—免疫学] R392.4[医药卫生—基础医学]