Detection of Antibodies in Blood Plasma Using Bioluminescent Sensor Proteins and a Smartphone
Detection of Antibodies in Blood Plasma Using Bioluminescent Sensor Proteins and a Smartphone
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DOI：
10.1021/acs.analchem.6b00534
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发表时间：
2016-04-19
影响因子：
7.4
通讯作者：
Merkx, Maarten
中科院分区：
文献类型：
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作者：
Arts, Remco;den Hartog, Ilona;Merkx, Maarten
抗体检测在许多诊断和生物分析测定中具有根本重要性，然而目前的检测技术往往是费力的和/或昂贵的。我们提出了一种基于生物发光共振能量转移（BRET）的新传感器平台（LUMABS），该平台允许使用智能手机作为唯一的设备直接检测溶液中的抗体。LUMABS是单蛋白传感器，其由通过半柔性接头连接到绿色荧光受体蛋白mNeon绿色的蓝光发射荧光素酶NanoLuc组成，所述荧光素酶NanoLuc使用辅助结构域保持在一起。抗体与接头侧翼的表位序列的结合破坏了辅助结构域之间的相互作用，导致BRET效率大幅降低。即使在皮摩尔浓度的抗体下，也可以在血浆中直接检测到所发射的光的颜色从绿-蓝到蓝色的变化。此外，LUMABS的模块化结构允许通过表位序列的简单交换来改变靶特异性，如本文针对抗HIV 1-p17、血凝素（HA）和I型登革热病毒的抗体所证明的。灵敏的比率生物发光检测和LUMABS设计的固有模块性的组合提供了一个有吸引力的通用平台，用于床旁抗体检测，避免了与常规免疫测定相关的复杂液体处理步骤。
Antibody detection is of fundamental importance in many diagnostic and bioanalytical assays, yet current detection techniques tend to be laborious and/or expensive. We present a new sensor platform (LUMABS) based on bioluminescence resonance energy transfer (BRET) that allows detection of antibodies directly in solution using a smartphone as the sole piece of equipment. LUMABS are single-protein sensors that consist of the blue-light emitting luciferase NanoLuc connected via a semiflexible linker to the green fluorescent acceptor protein mNeonGreen, which are kept close together using helper domains. Binding of an antibody to epitope sequences flanking the linker disrupts the interaction between the helper domains, resulting in a large decrease in BRET efficiency. The resulting change in color of the emitted light from, green-blue to blue can be detected directly in blood plasma, even at picomolar concentrations of antibody. Moreover, the modular architecture of LUMABS allows changing of target specificity by simple exchange of epitope sequences, as demonstrated here for antibodies against HIV1-p17, hemagglutinin (HA), and dengue virus type I. The combination of sensitive ratiometric bioluminescent detection and the intrinsic modularity of the LUMABS design provides an attractive generic platform for point-of-care antibody detection that avoids the complex liquid handling steps associated with conventional immunoassays.