The Single-Molecule Centroid Localization Algorithm Improves the Accuracy of Fluorescence Binding Assays,Biochemistry

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The Single-Molecule Centroid Localization Algorithm Improves the Accuracy of Fluorescence Binding Assays
Biochemistry ( IF 2.9 ) Pub Date : 2018-02-19 00:00:00 , DOI: 10.1021/acs.biochem.7b01293
Boyang Hua ₁ , Yanbo Wang ₁ , Seongjin Park ₂ , Kyu Young Han ₂ , Digvijay Singh ₁ , Jin H Kim ₃ , Wei Cheng ₃ , Taekjip Ha _{1,

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Affiliation

Here, we demonstrate that the use of the single-molecule centroid localization algorithm can improve the accuracy of fluorescence binding assays. Two major artifacts in this type of assay, i.e., nonspecific binding events and optically overlapping receptors, can be detected and corrected during analysis. The effectiveness of our method was confirmed by measuring two weak biomolecular interactions, the interaction between the B1 domain of streptococcal protein G and immunoglobulin G and the interaction between double-stranded DNA and the Cas9–RNA complex with limited sequence matches. This analysis routine requires little modification to common experimental protocols, making it readily applicable to existing data and future experiments.

中文翻译：

单分子质心定位算法提高了荧光结合测定的准确性

在这里，我们证明使用单分子质心定位算法可以提高荧光结合测定的准确性。此类测定中的两个主要伪影，即非特异性结合事件和光学重叠受体，可以在分析过程中检测和校正。通过测量两种微弱的生物分子相互作用，即链球菌蛋白 G 的 B1 结构域和免疫球蛋白 G 之间的相互作用，以及双链 DNA 和具有有限序列匹配的 Cas9-RNA 复合物之间的相互作用，证实了我们方法的有效性。该分析例程几乎不需要对常见实验方案进行修改，因此可以轻松应用于现有数据和未来的实验。

更新日期：2018-02-19

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