Protein sensors combining both on-and-off model for antibody homogeneous assay

• Published in: Biosensors & bioelectronics Vol. 209; p. 114226
• Main Authors: Li, Jie, Wang, Jin-Lan, Zhang, Wen-Lu, Tu, Zeng, Cai, Xue-Fei, Wang, Yu-Wei, Gan, Chun-Yang, Deng, Hai-Jun, Cui, Jing, Shu, Zhao-Che, Long, Quan-Xin, Chen, Juan, Tang, Ni, Hu, Xue, Huang, Ai-Long, Hu, Jie-Li
• Format: Journal Article
• Language: English
• Published: England Elsevier B.V 01.08.2022
• Subjects: antibodies; Antibodies, Viral; Antibody test; Biosensing Techniques; Biosensor; biosensors; blood serum; COVID-19 - diagnosis; enzyme activity; Hepatitis C virus; Homogeneous assay; Human immunodeficiency virus; Humans; luciferase; Luciferases; NanoLuc; Protein switch; SARS-CoV-2; signal-to-noise ratio; Homogeneous assay; NanoLuc; Biosensor; Protein switch; Antibody test
• ISSN: 0956-5663; 1873-4235; 1873-4235
• DOI: 10.1016/j.bios.2022.114226

Protein sensors based on allosteric enzymes responding to target binding with rapid changes in enzymatic activity are potential tools for homogeneous assays. However, a high signal-to-noise ratio (S/N) is difficult to achieve in their construction. A high S/N is critical to discriminate signals from the background, a phenomenon that might largely vary among serum samples from different individuals. Herein, based on the modularized luciferase NanoLuc, we designed a novel biosensor called NanoSwitch. This sensor allows direct detection of antibodies in 1 μl serum in 45 min without washing steps. In the detection of Flag and HA antibodies, NanoSwitches respond to antibodies with S/N ratios of 33-fold and 42-fold, respectively. Further, we constructed a NanoSwitch for detecting SARS-CoV-2-specific antibodies, which showed over 200-fold S/N in serum samples. High S/N was achieved by a new working model, combining the turn-off of the sensor with human serum albumin and turn-on with a specific antibody. Also, we constructed NanoSwitches for detecting antibodies against the core protein of hepatitis C virus (HCV) and gp41 of the human immunodeficiency virus (HIV). Interestingly, these sensors demonstrated a high S/N and good performance in the assays of clinical samples; this was partly attributed to the combination of off-and-on models. In summary, we provide a novel type of protein sensor and a working model that potentially guides new sensor design with better performance. [Display omitted]