一种基于酶联免疫吸附测定的结合与竞争方法，用于快速确定配体-受体相互作用。
An ELISA Based Binding and Competition Method to Rapidly Determine Ligand-receptor Interactions.
作者信息
Syedbasha Mohameedyaseen, Linnik Janina, Santer Deanna, O'Shea Daire, Barakat Khaled, Joyce Michael, Khanna Nina, Tyrrell D Lorne, Houghton Michael, Egli Adrian
机构信息
Applied Microbiology Research, Department of Biomedicine, University of Basel.
Applied Microbiology Research, Department of Biomedicine, University of Basel; Department of Biosystems Science and Engineering, ETH Zurich, and Swiss Institute of Bioinformatics; Swiss Institute of Bioinformatics.
出版信息
J Vis Exp. 2016 Mar 14(109):53575. doi: 10.3791/53575.
A comprehensive understanding of signaling pathways requires detailed knowledge regarding ligand-receptor interaction. This article describes two fast and reliable point-by-point protocols of enzyme-linked immunosorbent assays (ELISAs) for the investigation of ligand-receptor interactions: the direct ligand-receptor interaction assay (LRA) and the competition LRA. As a case study, the ELISA based analysis of the interaction between different lambda interferons (IFNLs) and the alpha subunit of their receptor (IL28RA) is presented: the direct LRA is used for the determination of dissociation constants (KD values) between receptor and IFN ligands, and the competition LRA for the determination of the inhibitory capacity of an oligopeptide, which was designed to compete with the IFNLs at their receptor binding site. Analytical steps to estimate KD and half maximal inhibitory concentration (IC50) values are described. Finally, the discussion highlights advantages and disadvantages of the presented method and how the results enable a better molecular understanding of ligand-receptor interactions.
全面了解信号通路需要有关配体 - 受体相互作用的详细知识。本文介绍了两种用于研究配体 - 受体相互作用的快速且可靠的逐点酶联免疫吸附测定（ELISA）方案：直接配体 - 受体相互作用测定（LRA）和竞争性LRA。作为案例研究，展示了基于ELISA对不同λ干扰素（IFNLs）与其受体α亚基（IL28RA）之间相互作用的分析：直接LRA用于测定受体与IFN配体之间的解离常数（KD值），竞争性LRA用于测定一种寡肽的抑制能力，该寡肽旨在在其受体结合位点与IFNLs竞争。描述了估计KD和半数最大抑制浓度（IC50）值的分析步骤。最后，讨论突出了所提出方法的优缺点，以及这些结果如何有助于更好地从分子层面理解配体 - 受体相互作用。
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