Powers Glenn A, Hudson Peter J, Wheatcroft Michael P
Avipep Pty Ltd, Parkville, Australia.
Methods Mol Biol. 2012;907:699-712. doi: 10.1007/978-1-61779-974-7_39.
Multimeric antibody fragments, particularly dimers (diabodies), trimers (triabodies), and tetramers (tetrabodies) of single-chain Fv molecules (scFv), provide high avidity through multivalent binding to the target antigen. The combination of their smaller size and avid binding can provide desirable biological characteristics for tumor targeting applications in vivo; for example, diabodies can have greater tumor penetration and faster blood clearance rates compared to intact full-size antibodies (IgGs). The pharmacokinetic and biodistribution characteristics can further be optimized by the addition of specific thiolation sites for conjugation of PEG molecules to regulate molecular weight and reduce kidney uptake. Thiolation sites can also be used for precise loading of therapeutic payloads. This protocol describes our method for construction and bacterial production of soluble multimeric antibody scFv fragments, focusing on diabodies (scFv dimers).
多聚体抗体片段，特别是单链Fv分子（scFv）的二聚体（双特异性抗体）、三聚体（三特异性抗体）和四聚体（四特异性抗体），通过与靶抗原的多价结合提供高亲和力。它们较小的尺寸和高亲和力结合的组合可为体内肿瘤靶向应用提供理想的生物学特性；例如，与完整的全尺寸抗体（IgG）相比，双特异性抗体可具有更高的肿瘤穿透性和更快的血液清除率。通过添加特定的巯基化位点以与PEG分子偶联来调节分子量并减少肾脏摄取，可进一步优化药代动力学和生物分布特性。巯基化位点还可用于精确加载治疗性有效载荷。本方案描述了我们构建和细菌生产可溶性多聚体抗体scFv片段的方法，重点是双特异性抗体（scFv二聚体）。
