RAPPID-M:一种基于通用抗体结合蛋白M的混合测量生物发光三明治免疫分析法。

IF 9.1 1区 化学 Q1 CHEMISTRY, ANALYTICAL
Anna Swietlikowska,Eva van Aalen,Max Bossink,Laura van Weesep,Maarten Merkx
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引用次数: 0

摘要

均质免疫分析允许直接在样品中检测生物标志物,为需要多次洗涤和孵育步骤的传统异质免疫分析提供了有吸引力的替代方案。我们之前报道了RAPPID的发展,RAPPID是一种生物发光传感器平台,它使用分裂荧光素酶互补来直接在溶液中进行三明治免疫测定。尽管RAPPID比传统的异质免疫测定法提供了许多好处,但它需要蛋白g介导的荧光素酶片段与IgG单克隆抗体Fc部分的光偶联。在这里,我们报道了新一代RAPPID传感器(RAPPID-M),它不依赖于蛋白质G的光偶联,而是使用来自蛋白质M的通用抗体结合域。蛋白质M的使用大大扩展了RAPPID的应用范围,包括蛋白质G不相容但在分析上重要的小鼠IgG1抗体,以及scFv和Fab抗体片段。此外,发现蛋白质M的结合基本上是不可逆的,取消了光交联的需要。RAPPID- m在原有RAPPID平台的基础上进行了改进,通过抗体-分裂-荧光素酶偶联物的混合-测量型组装,并与更广泛的抗体类型兼容,而不影响甚至提高了分析性能。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
RAPPID-M: A Mix-and-Measure Bioluminescent Sandwich Immunoassay Based on Generic Antibody-Binding Protein M.
Homogeneous immunoassays that allow direct in-sample detection of biomarkers provide an attractive alternative to traditional heterogeneous immunoassays that require multiple washing and incubation steps. We previously reported the development of RAPPID, a bioluminescent sensor platform that uses split luciferase complementation to enable sandwich immunoassays directly in solution. Although RAPPID provides many benefits over traditional heterogeneous immunoassays, it requires protein G-mediated photoconjugation of split luciferase fragments to the Fc part of IgG monoclonal antibodies. Here, we report a new generation of RAPPID sensors (RAPPID-M) that do not rely on photoconjugation of protein G, but instead use a generic antibody-binding domain derived from protein M. The use of protein M substantially expands the application of RAPPID to include protein G-incompatible but analytically important mouse IgG1 antibodies, as well as scFv and Fab antibody fragments. Moreover, binding of protein M is found to be essentially irreversible, abolishing the need for photo-cross-linking. RAPPID-M thus improves upon the original RAPPID platform by the mix-and-measure-type assembly of antibody-split-luciferase conjugates and its compatibility with a broader scope of antibody types without compromising and even improving the analytical performance.
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来源期刊
ACS Sensors
ACS Sensors Chemical Engineering-Bioengineering
CiteScore
14.50
自引率
3.40%
发文量
372
期刊介绍: ACS Sensors is a peer-reviewed research journal that focuses on the dissemination of new and original knowledge in the field of sensor science, particularly those that selectively sense chemical or biological species or processes. The journal covers a broad range of topics, including but not limited to biosensors, chemical sensors, gas sensors, intracellular sensors, single molecule sensors, cell chips, and microfluidic devices. It aims to publish articles that address conceptual advances in sensing technology applicable to various types of analytes or application papers that report on the use of existing sensing concepts in new ways or for new analytes.
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