噬菌体模拟抗原替代亲环素A在测定自身抗体中的应用

目的 探讨噬菌体展示肽库来源的模拟抗原替代重组人亲环素A(rhCyPA)在测定自身免疫病人血清抗亲环素自身抗体中应用价值。方法 以抗人亲环素A单克隆抗体(McAb)D4和E6作捕获抗体对噬菌体展示肽库进行淘筛，通过序列测定推断出模拟抗原氨基酸序列。再以筛出的模拟抗原包被微孔板，与重组人亲环素A抗原进行对比，用ELJSA法检测系统性红斑狼疮患者血清抗亲环素自身抗体。结果 从12肽库淘筛出的10个噬菌体克隆中9个共有一致氨基酸序列HWEYTISPHPRL。56例SLE患者血清用12肽模拟抗原的ELISA测定，CyPA自身抗体阳性率为50％(28／56)，用rhCyPA作包被抗原测定阳性率为43％(214／56)；两法检测34例其他自身免疫病的阳性率均为11％，两法结果符合率为95．6％。通过试验比较，12肽模拟抗原用于检测CyPA自身抗体优于重组抗原和7肽模拟抗原。结论用抗CyPA单克隆抗体从噬菌体展示肽库淘筛出的特异12肽能模拟亲环素A抗原表位信息，可以代替亲环素A用来检测其自身抗体。

Using mimicry epitopes from phage displayed peptide library as substitute antigen for cyclophilin A in ELISA for detection of anti-CyPA autoantibody

Objective To use mimicry epitopes from phage displayed peptide library as substitute for recombinant human cyclophilin A (rhCyPA) in detection of CyPA autoantibody in sera of patients with systemic lupus erythematosus (SLE).Methods Monoclonal antibody D4 and E6 were used as the capture antibody; the Ph.D.-12 p hage display peptide library was screened by biopanning. Ten phage clones with h igh affinity for McAb D4 and E6 (five for each) were chosen. Their inserts were sequenced. The amino sequence of mimicry epitope was subsequently reasoned out. Selected 12 peptide and previously obtained 7 peptide mimicry epitopes were used as coated antigen and compared with rhCyPA in ELISA for anti-CyPA autoantibody .Results Nine of ten phage clones with high affinity for McAb D4 and E6 (five for D4, four for E6) shared a consensus amino sequence HW EYTTSPHPRL. In comparative study of ELISA, 56 serum samples from SLE patients we re tested for anti-CyPA autoantibody, 28 samples were positive (50%) with 12 pe ptide mimicry antigen and 24 were positive (43%) with rhCyPA, 4 of 34 (11%) samp les from those with other autoimmune disorders were positive with both antigens, the coincidence rate between two methods was 95 6%. The research also showed t hat 12 peptide mimicry antigen has the advantages of rhCyPA and 7 peptide mimicr y antigen in detecting anti-CyPA autoantibody. Conclusions The specific 12 peptide screened from phage-displayed peptide library by ant i-CyPA monoclonal antibody can mimic epitope information of cyclophilin A well and be used as substitute for rhCyPA in ELISA.