Chu Xiao Xia, Hou Ming, Peng Jun, Zhu Yuan Yuan, Ji Xue Bin, Wang Lin, Zhang Feng, Ma Dao Xin
Hematology Oncology Center, Qilu Hospital of Shandong University, Jinan, Shandong, China.
Eur J Haematol. 2006 Feb;76(2):153-9. doi: 10.1111/j.1600-0609.2005.00557.x.
To make humanized monoclonal antibodies by phage surface display technology, we screened out the specific anti-platelet glycoproteins (GPs) IgG antibody from patients with chronic idiopathic thrombocytopenic purpura (ITP), which can inhibit platelet aggregation.
We studied plasmas from 68 patients with ITP for the presence of IgG antibodies specific for GPIIb/IIIa and/or GPIb/IX using modified monoclonal antibody immobilization of platelet antigen assays. The IgG antibody and its F(ab')(2) fragments of the positive plasmas which could inhibit platelet aggregation function were prepared and purified. Their immunoreactivity to platelet GPs and effects on platelet function were further analyzed.
GPIIb/IIIa- and GPIb/IX-specific antibodies were found in 21 and 19 patients, respectively. Six of them had antibodies against both GP complexes. Among the 34 positive plasmas, four with positive anti-GPIIb/IIIa autoantibody showed significant inhibition of platelet aggregation induced by adenosine diphosphate (ADP), whereas one with GPIb/IX-specific antibody inhibited ristocetin-induced platelet aggregation. The purified IgG and its F(ab')(2) fragments from two patients not only retained the ability to bind to platelet GPs but also impaired the in vitro ADP-induced platelet aggregation.
F(ab')(2) portion of the IgG is a functional fragment, which is responsible for the autoantibody interaction with platelet GPs in ITP, and some of them also affect platelet function, which can be used to develop completely humanized anti-GPIIb/IIIa small molecular phage antibody.
为利用噬菌体表面展示技术制备人源化单克隆抗体，我们从慢性特发性血小板减少性紫癜（ITP）患者中筛选出能抑制血小板聚集的特异性抗血小板糖蛋白（GPs）IgG抗体。
我们采用改良的血小板抗原单克隆抗体固定试验，研究了68例ITP患者血浆中针对GPIIb/IIIa和/或GPIb/IX的IgG抗体的存在情况。制备并纯化了能抑制血小板聚集功能的阳性血浆中的IgG抗体及其F(ab')(2)片段。进一步分析了它们与血小板GPs的免疫反应性及对血小板功能的影响。
分别在21例和19例患者中发现了GPIIb/IIIa特异性抗体和GPIb/IX特异性抗体。其中6例同时具有针对两种GP复合物的抗体。在34份阳性血浆中，4份抗GPIIb/IIIa自身抗体阳性的血浆对二磷酸腺苷（ADP）诱导的血小板聚集有显著抑制作用，而1份GPIb/IX特异性抗体阳性的血浆抑制了瑞斯托霉素诱导的血小板聚集。从2例患者中纯化的IgG及其F(ab')(2)片段不仅保留了与血小板GPs结合的能力，还损害了体外ADP诱导的血小板聚集。
IgG的F(ab')(2)部分是一个功能性片段，它在ITP中负责自身抗体与血小板GPs的相互作用，其中一些还影响血小板功能，可用于开发完全人源化的抗GPIIb/IIIa小分子噬菌体抗体。
