[Antibody engineering-based approach for hapten immunometric assays with high sensitivity].
[Antibody engineering-based approach for hapten immunometric assays with high sensitivity].
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DOI：
10.1248/yakushi.127.55
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发表时间：
2007
期刊：
影响因子：
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通讯作者：
N. Kobayashi;Y. Kato;H. Oyama;J. Goto
中科院分区：
文献类型：
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作者：
N. Kobayashi;Y. Kato;H. Oyama;J. Goto
低分子量生理活性物质的痕量表征（例如，类固醇、儿茶酚胺、肾上腺素和寡肽）是临床分析中的重要对象，竞争性免疫测定通常用于此目的。然而，半抗原的亚飞摩尔范围的测定是非常困难的，因为竞争性免疫测定的灵敏度基本上受到抗半抗原抗体的亲和力的限制，所述亲和力几乎没有达到10（11）（l/mol）的范围作为亲和力常数（K（α））。虽然非竞争性的"免疫测定"形式，双位点免疫测定（夹心免疫测定），甚至能够对大分子如蛋白质进行亚阿摩尔范围的测量，但该原理不能直接应用于半抗原，因为它们的低分子量阻止了两个抗体分子的同时结合。为了克服这些限制，我们需要通过抗体分子的蛋白质工程（"抗体工程"）或建立适用于半抗原的新型免疫测定形式来产生显示对半抗原的亲和力的人工抗体。本文综述了亚飞摩尔范围内半抗原测定的背景和最新方法。我们的研究发展的半抗原免疫测定也被描述。
The trace characterization of physiologically active substances with low molecular weight (e.g., steroids, catecholamines, prostaglandins, and oligopeptides), which are classified as "haptens", is an important subject in clinical analysis, and competitive immunoassays have conventionally been used for this purpose. However, the subfemtomole-range determination of haptens is very difficult, as the sensitivity of competitive immunoassays is essentially limited by the affinity of the anti-hapten antibodies that barely reaches the range of 10(11) (l/mol) as the affinity constant (K(a)). Although a noncompetitive "immunometric assay" format, the two-site immunometric assay (sandwich immunoassay), enables even subattomole-range measurements of macromolecules such as proteins, this principle can not be directly applied to haptens, as their low molecular mass prohibits simultaneous binding by two antibody molecules. To overcome such limitations, we are required either to create artificial antibodies showing ultrahigh affinity to haptens by protein engineering of antibody molecules ("antibody engineering") or establishment of novel immunometric assay formats applicable to haptens. This review surveys the background and recent approach for subfemtomole-range determination of haptens using novel immunometric assay methods. Our studies for the development of hapten immunometric assays are also described.