Monoclonal antibody binding-site diversity assessment with a cell-based clustering assay
Monoclonal antibody binding-site diversity assessment with a cell-based clustering assay
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DOI：
10.1016/j.jim.2013.12.007
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发表时间：
2014-03-01
影响因子：
2.2
通讯作者：
Theunissen, Jan-Willem
中科院分区：
文献类型：
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作者：
Liao-Chan, Sindy;Zachwieja, Joseph;Theunissen, Jan-Willem
靶向特定抗原的一组抗体的多样性可以以各种测定形式建立。在常规表位分箱测定中，以成对方式测试纯化的抗体：将彼此竞争结合抗原的两种抗体分组到相同的簇或箱中，而当它们不竞争时，将它们分配到两个不同的簇。在这里，我们提出了一种高通量测定法，该测定法能够在不需要抗体纯化的情况下对粗杂交瘤上清液进行分组。此外，该测定不需要重组蛋白，因为它是在表达目标抗原的细胞上进行的。因此，对于不适合纯化的细胞表面蛋白，可以使用抗体聚类测定。重链可变区（V-H）测序显示簇内的VH组成是保守的。最后，该测定与使用纯化抗原的常规表位分箱测定具有良好的一致性。(C)2013爱思唯尔有限公司版权所有。
The diversity of a panel of antibodies that target a specific antigen can be established in various assay formats. In conventional epitope binning assays purified antibodies are tested in a pairwise manner: two antibodies that compete with each other for binding to an antigen are grouped into the same cluster or bin, while they are assigned to two different clusters when they do not compete. Here we present a high through put assay that enables grouping of crude hybridoma supernatants without a need for antibody purification. In addition, the assay does not require recombinant protein, because it is conducted on cells that express the antigen of interest. Hence, one can use the antibody-clustering assay for cell surface proteins that are not amenable to purification. Heavy chain variable region (V-H) sequencing shows that VH composition within clusters is conserved. Finally, the assay is in good agreement with a conventional epitope binning assay with purified antigen. (C) 2013 Elsevier B.V. All rights reserved.