Title | Highly Sensitive and Specific Detection of Influenza A Viruses Using Bimolecular Fluorescence Complementation (BiFC) Reporter System. | ||
Author | Lee, Ui Jin; Oh, Yunkwang; Kwon, Oh Seok; Shin, Yong-Beom; Kim, Moonil | ||
Journal | Biosensors (Basel) | Publication Year/Month | 2023-Aug |
PMID | 37622868 | PMCID | PMC10452828 |
Affiliation + expend | 1.Critical Diseases Diagnostics Convergence Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahang-ro, Yuseong-gu, Daejeon 34141, Republic of Korea. |
In this study, we developed a highly sensitive and specific bimolecular fluorescence complementation (BiFC)-based influenza A virus (IAV)-sensing system by combining a galactose/glucose-binding protein (GGBP) with an N-terminal large domain (YN1-172) and a C-terminal small domain (YC173-239) made up of enhanced yellow fluorescence protein (eYFP). The GGBP-based BiFC reporter exhibits the fluorescence reconstitution as a result of conformational changes in GGBP when lactose, which was derived from 6\'-silalyllactose and used as a substrate for neuraminidase (NA), binds to GGBP in the presence of IAV. The system showed a linear dynamic range extending from 1 x 10(0) to 1 x 10(7) TCID(50)/mL, and it had a detection limit of 1.1 x 10(0) TCID(50)/mL for IAV (H1N1), demonstrating ultra-high sensitivity. Our system exhibited fluorescence intensity enhancements in the presence of IAV, while it displayed weak fluorescence signals when exposed to NA-deficient viruses, such as RSV A, RSV B, adenovirus and rhinovirus, thereby indicating selective responses for IAV detection. Overall, our system provides a simple, highly sensitive and specific IAV detection platform based on BiFC that is capable of detecting ligand-induced protein conformational changes, obviating the need for virus culture or RNA extraction processes.