EGFR特异性单链抗体与靶细胞结合的特异性

The Specificity of EGFR scFv Binding with Target Cells

目的:检测EGFR特异性单链抗体F4-scFv与EGFR阳性及阴性细胞结合特性。方法:提取细菌周质腔蛋白经N i-NTA柱纯化F4-scFv;采用流式细胞术及W estern b lot检测F4-scFv与EGFR阳性及阴性细胞结合的特异性。结果:N i2+柱纯化的F4-scFv经SDS-PAGE可见分子量为36 kDa单一条带,F4-scFv可与EGFR阳性细胞CHO-EGFR-GFP1、A431及MDA-MB-231特异性结合,而不与EGFR阴性细胞CHO-K1及SKOV3结合。结论:F4-scFv可与天然表达EGFR的细胞特异性结合,为其靶向抗肿瘤研究奠定了基础。

Objective To study the binding properties of an EGFR scFv （F4 - scFv） to EGFR positive and negative cell lines. Methods F4 - scFv was purified from periplasmic fraction of bacteria by IMAC using a Ni - NTA agarose column ; The binding properties of F4 - scFv to EGFR positive and negative cells were analyzed by FCM and Western blot. Results The purified F4 - scFv was migrated as a single band in a SDS - PAGE analysis with molecular size of about 36kDa; F4 - scFv bound EGFR positive cell line CHO - EGFR - GFP1, A431, MDA - MB -231, but didnt bind EGFR negative cell line CHO - K1 and SKOV3. Conclusion F4 - scFv binds specifically to EGFR positive cell lines, this work provides a basis for further research of anti - tumor drugs using this scFv as target molecule.