Novel monoclonal antibodies to cyclosporine A: characterization and epitope mapping with cyclosporine analogs and cyclophilin.
Novel monoclonal antibodies to cyclosporine A: characterization and epitope mapping with cyclosporine analogs and cyclophilin.
复制标题
环孢素 A 的新型单克隆抗体：环孢素类似物和亲环蛋白的表征和表位作图。
DOI：
10.1016/0161-5890(92)90162-q
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发表时间：
1992
影响因子：
3.6
通讯作者：
Erlanger,BF
中科院分区：
文献类型：
--
作者：
Cacalano,NA;Aggarwal,R;Quesniaux,VF;Cleveland,WL;Erlanger,BF
用光敏交联剂4-苯甲酰基苯甲酸（BBa）将环孢素A（CsA）与蛋白质载体连接，制备了CsA单克隆抗体。以Cs-BBa-牛血清白蛋白（Cs-BBa-BSA）为免疫原，制备了22株抗Cs单克隆抗体。他们的特点是相对于亲和力的Scatchard分析的放射免疫测定法（RIA），和相对于特异性的ELISA，其中一系列的单取代Cs衍生物作为抑制剂进行了检查。单克隆抗体的亲和力范围为5 × 10 − 8 M至2 × 10 − 10 M，根据Cs类似物的ELISA抑制数据，以及与两个Cs分子相对侧暴露的Cs-BSA结合物的结合，抗体分为五个表位识别组。还通过ELISA与亲环蛋白（CyP）竞争研究了与Cs的结合，亲环蛋白是一种Cs结合蛋白，其表位特异性已被很好地表征。发现CyP的竞争与抗体特异性相关，而与亲和力无关，即CyP与特异性与CyP最相似的抗体竞争得最好。因此，表位作图可以在两种不同种类的结合蛋白竞争相同抗原的系统中完成。这种类型的表征可用于鉴定结合位点模拟受体结合位点的抗体。
Monoclonal antibodies to cyclosporine A (Cs), a potent immunosuppressant, were generated in BALB/c mice using a novel antigen prepared by linking Cs to a protein carrier via a photoactive cross-linking reagent, 4-benzoylbenzoic acid (BBa). Twenty-two monoclonal anti-Cs antibodies were generated, using Cs-BBa-bovine serum albumin (Cs-BBa-BSA) as the immunogen. They were characterized with respect to affinity by Scatchard analysis of a radioimmunoassay (RIA), and with respect to specificity by an ELISA in which a series of singly substituted Cs derivatives were examined as inhibitors. McAb affinities ranged from 5 × 10−8M to 2 × 10−10M, Based on ELISA inhibition data with Cs analogs, and on the binding to two Cs-BSA conjugates in which opposite sides of the Cs molecule are exposed, the antibodies fell into five epitope recognition groups. Binding to Cs was also studied by ELISA in competition with cyclophilin (CyP), a Cs-binding protein whose epitope specificity has been well characterized. Competition by CyP was found to correlate with antibody specificity, not with affinity, i.e. CyP competed best with antibodies having specificities most similar to that of CyP. Epitope mapping can, therefore, be accomplished in a system in which two different species of binding proteins compete for the same antigen. This type of characterization may be useful in identifying antibodies whose combining sites mimic those of a receptor.