Hybrid Cell Formation by Targeted Electrofusion

产生永生抗体的杂交瘤通过以下融合产生： 产生抗体的脾细胞与骨髓瘤细胞。广泛使用的 化学聚变过程是时间密集的，需要大量的 并提供相对低产量的所需抗原 杂交瘤 传统的化学聚变会导致 杂交瘤的产生，其中大多数不产生所需的 抗体的 这项建议旨在制定方法， 这将为细胞融合过程增加抗原特异性。 骨髓瘤 首先用抗原（或半抗原）包被细胞。 这些被标记的细胞 然后与来自免疫小鼠的脾细胞一起孵育， 它们与互补抗体结合。两种细胞之间的融合可以 然后通过化学或电融合来引发。 结果 第一阶段项目表明：1）可以标记 骨髓细胞表面的半抗原或抗原，2）在适当的 3）在与脾孵育后， 细胞，表面标记的骨髓瘤细胞可以找到互补的淋巴细胞 通过分子细胞选择，和4）靶向骨髓瘤。淋巴细胞 二聚体可以融合以产生杂交瘤。 还证明了 高速流式细胞术提供了一种灵敏的监测方法 这些过程。现在已经证明了可行性，第二阶段 将提供支持，以调查生产的最佳条件 通过靶向电融合的杂交瘤。 PI希望利用靶向细胞电融合技术， 更高效的杂交瘤生产。 常规化学细胞融合 导致杂交瘤的随机产生，其中大多数不是 产生所需的抗体。 PI在这里建议利用 B淋巴细胞的抗原结合特性，以增加特异性 细胞融合的过程。这将克服一个重要的限制 到定制杂交瘤生产的商业化。

Immortal antibody producing hybridomas are produced by the fusion of antibody producing spleen cells with myeloma cells. The widely used chemical fusion procedures are time intensive, require large amounts of valuable antigen and provide relatively low yields of the desired hybridomas. Conventional chemical fusion results in the random generation of hybridomas, most of which are not producing the desired antibodies. This proposal is directed towards the development of methods which will add antigen specificity to the cell fusion process. Myeloma cells are first coated with the antigen (or hapten). These labeled cells are then incubated with spleen cells from an immunized mouse, whereupon they bind to complementary antibody. Fusion between the two cells can then be intiated either chemically or by electrofusion. Results from the Phase I project have indicated that: 1) It is possible to label the surface of myelome cells with haptens or antigens, 2) Under proper conditions, the labeled cells are viable, 3) Upon incubation with spleen cells, surface labeled myeloma cells can find complementary lymphocytes by molecular cell selection, and 4) The targeted myeloma.lymphocyte dimers can be fused to yield hybridomas. It was also demonstrated that high speed flow cytometry provides a sensitive method for monitoring these processes. Now that feasibility has been demonstrated, Phase II would provide support to investigate optimal conditions for producing hybridomas via targeted electrofusion. The PI wishes to exploit the technique of targeted cell electrofusion for more efficient hybridoma production. Conventional chemical cell fusions result in random generation of hybridomas, most of which are not producing the desired antibody. The PI here proposes to exploit the antigen.binding properties of B.lymphocytes to increase the specificity of the cell fusion process. This would overcome an important limitation to the commercialization of custom hybridoma production.