Properties of two monoclonal antibodies directed against the Fc and Fab' regions of rat IgE.
Properties of two monoclonal antibodies directed against the Fc and Fab' regions of rat IgE.
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针对大鼠 IgE Fc 和 Fab 区域的两种单克隆抗体的特性。
DOI：
10.1159/000233347
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发表时间：
1983
期刊：
影响因子：
--
通讯作者：
Mohanakumar,T
中科院分区：
文献类型：
--
作者：
Conrad,DH;Studer,E;Gervasoni,J;Mohanakumar,T
通过用大鼠IgE免疫BALB/c小鼠并将脾细胞与非分泌性浆细胞瘤P3/X63Ag8.653融合来产生针对大鼠IgE的Fc和Fab部分的杂交瘤抗体。其中两种抗体（称为 A2 和 B5）已得到广泛表征。使用来自 IR 162 和 IR 2 免疫细胞瘤的大鼠 IgE 和大鼠 IgG 进行的竞争性结合实验表明，A2 和 B5 都是ε链特异性的，而不是抗独特型的。 A2 还表现出与小鼠 IgE 的一些交叉反应性。当用胰凝乳蛋白酶处理IgE以产生F(ab')2和Fab片段两者时，酶处理的IgE保留了与B5的反应性，但失去了与A2的反应性。 IgE 在 56 °C 下热变性导致 IgE 对大鼠嗜碱性白血病细胞上 A2 和 Fc 受体的反应性逐渐丧失； B5的反应活性保持不变。 A2 与 IgE Fc 上参与与大鼠嗜碱性白血病细胞 Fc 受体结合的相同位点没有明显相互作用； A2对IgE与大鼠嗜碱性白血病细胞的结合几乎没有影响。因此，数据表明B5的抗原位点位于IgE分子的Fab区并且A2与IgE Fc反应。在双标记实验中还评估了使用这些抗体测量细胞结合的 IgE，并讨论了使用二价抗体定量细胞表面抗原的潜在问题。
Hybridoma antibodes directed against the Fc and Fab portions of rat IgE were produced by immunizing BALB/c mice with rat IgE and fusing the spleen cells with the nonsecreting plasmacytoma P3/X63Ag8.653. Two of the antibodies, designated as A2 and B5, were extensively characterized. Competitive binding experiments using rat IgE from the IR 162 and IR 2 immunocytomas and rat IgG indicated that both A2 and B5 wereε-chain specific and not anti-idiotype. A2 also exhibited some cross-reactivity with mouse IgE. When IgE was treated with chymotrypsin so as to produce both F(ab′)2and Fab fragments, the enzyme-treated IgE retained reactivity with B5, but the reactivity to A2 was lost. Heat denaturation of IgE at 56 °C resulted in a progressive loss of reactivity of the IgE for both A2 and the Fc receptor on rat basophilic leukemia cells; the reactivity of B5 remained unchanged. A2 does not evidently interact with the same site on the Fc of IgE that is involved in binding to the rat basophilic leukemia cell Fc receptor; A2 exerted little influence on the binding of IgE to rat basophilic leukemia cells. Thus, the data indicate that the antigenic site for B5 is in the Fab region of the IgE molecule and that A2 reacts with the IgE Fc. Use of these antibodies to measure cell-bound IgE was also evaluated in dual label experiments, and potential problems in using divalent antibodies to quantitate cell surface antigens are discussed.