Development and application of CK-MB specific monoclonal antibodies
- 生命科学－已发表论文 
本研究拟建立肌酸激酶同工酶MB(CK-MB)特异性单克隆抗体(mAb)的研制方法,对抗CK-MB单抗进行评价分类及性质鉴定,并初步建立CK-MB; 定量检测试剂。以CK-MB抗原免疫BALB/c小鼠,利用常规单抗制备技术,使用间接和捕获ELISA差异筛选法筛选单抗。利用肌酸激酶同工酶(CK-; MM/BB/MB)抗原对所制备单抗的抗原识别表位进行鉴定,另通过免疫印迹法及合成CK-MM、CK-BB差异性的线性表位肽鉴定对所制备的单抗进行评; 价分类。使用双抗体夹心ELISA方法筛选检测CK-MB抗原的配对mAb,并初步建立CK-MB定量检测试剂。使用74例临床标本初步评价该试剂与罗氏; 试剂的检测一致性。最终,我们成功筛选到22株稳定分泌抗CK-MB抗体的杂交瘤细胞株,这些单抗可以分为线性、偏构象的CK-MB和CK-MM或者CK; -BB交叉的单抗以及与CK-MB特异反应的偏构象型单抗,并使用偏构象型单抗研制出CK-MB定量检测试剂,该试剂与罗氏试剂相关系数r达到0.930; 9。综上所述,本研究建立了研制CK-MB偏构象型特异性单抗的筛选方法,通过对所筛选的单抗进行分析鉴定并建立了CK-MB定量检测试剂,与罗氏试剂检; 测结果符合率高。The aim of this study is to develop creatine kinase isoenzyme MB (CK-MB); specific monoclonal antibodies (mAb), and characterize the monoclonal; antibody and further development of quantitative detection assay for; CK-MB. The BALB/c mice were immunized with purchased CK-MB antigen, then; monoclonal antibodies were prepared according to conventional hybridoma; technique and screened by indirect and capture ELISA method. To identify; the epitopes and evaluate the classification, purchased creatine kinase; isoenzyme MB (CK-MM/BB/MB) antigen was used to identify the epitopes,; with immunoblotting and synthetic CK-MM and CK-BB in different linear; epitope. A double antibody sandwich ELISA was applied to screen the mAb; pairs for CK-MB detection, and the quantitative detection assay for; CK-MB was developed. We used 74 cases of clinical specimens for; comparison of our assay with Roche's CK-MB assay. We successfully; developed 22 strains of hybridoms against CK-MB, these mAbs can be; divided into linear, partial conformational CK-MB, CK-MM or CK-BB cross; monoclonal antibody and CK-MB specific reaction with partial; conformational monoclonal antibody, and CK-MB quantitative detection; assay was developed by using partial conformational monoclonal antibody.; The correlation coefficient factor r of our reagent and Roche's was; 0.930 9. This study established a screening method for CK-MB partial; conformational specific monoclonal antibody, and these monoclonal; antibodies were analyzed and an established quantitative detection assay; was developed. The new assay had a high concordance with Roche's.