A Genetically Encoded Bioluminescent System for Fast and Highly Sensitive Detection of Antibodies with a Bright Green Fluorescent Protein
A Genetically Encoded Bioluminescent System for Fast and Highly Sensitive Detection of Antibodies with a Bright Green Fluorescent Protein
复制标题
一种基因编码生物发光系统，可通过亮绿色荧光蛋白快速、高灵敏地检测抗体
DOI：
10.1021/acsnano.1c05164
复制
发表时间：
2021-11-23
期刊：
影响因子：
17.1
通讯作者：
Chu, Jun
中科院分区：
文献类型：
--
作者：
Deng, Mengying;Yuan, Jing;Chu, Jun
一种快速、高灵敏的血清抗体检测方法将大大有助于疾病和感染的早期诊断和治疗性抗体的剂量优化。基于蓝色荧光素酶Nluc和绿色荧光蛋白(FP)mNeonGreen之间的生物发光共振能量转移(BRET)，LUMABS(更名为mNeonG-LUMABS，其中mNeonG是mNeonGreen的缩写)传感器的生物发光检测已被证明能够以合理的灵敏度快速检测血清中的抗体。然而，一些mNeonG-LUMABS传感器表现出较低的灵敏度，因此，灵敏度的提高仍然势在必行。在这里，我们报道了一种明亮的绿色FP，Clover4，它是通过对绿色FP三叶草进行结构引导突变而获得的。尽管三叶草和mNeonGreen的亮度和荧光光谱相似，但Clover4-LUMABS传感器的动态范围(最大20倍)和检测下限(LOD)要低得多(最大5.6倍)，这很可能是因为Clover4位于LUMABS中与Nluc更平行的方向。由于模块化设计，Clover4-LUMABS提供了一个通用的BRET系统，用于快速和高灵敏的血清抗体检测。
A method for fast and highly sensitive detection of antibodies in serum would greatly facilitate the early diagnosis of disease and infection and dose optimization of therapeutic antibody. Bioluminescence detection with LUMABS (renamed mNeonG-LUMABS, where mNeonG is short for mNeonGreen) sensors based on bioluminescence resonance energy transfer (BRET) between blue-emitting luciferase Nluc and green fluorescent protein (FP) mNeonGreen has been demonstrated to enable fast detection of antibodies directly in serum with reasonable sensitivity. However, some mNeonG-LUMABS sensors exhibit low sensitivity, and thus, sensitivity improvement remains imperative. Here, we report a bright green FP, Clover4, obtained by structure-guided mutagenesis of green FP Clover. Despite similar brightness and fluorescence spectra of Clover and mNeonGreen, Clover4-LUMABS sensors exhibit a largely increased dynamic range (maximum 20-fold) and much lower limit of detection (LOD) (maximum 5.6-fold), most likely because Clover4 is positioned in a more parallel orientation to Nluc in LUMABS. Due to modular design, Clover4-LUMABS offers a general BRET system for fast and highly sensitive antibody detection in serum.