Bioluminescent label-free molecular switch for SARS-CoV-2 detection

Area of Science:

• Biotechnology
• Molecular Biology
• Biochemistry

Background:

• Simple, single-step platforms are crucial for advancing diagnostic testing beyond traditional laboratory settings.
• Molecular switch platforms offer a promising strategy for virus detection due to their modularity, sensitivity, and adaptability.

Purpose of the Study:

• To develop a novel one-step bioluminescent molecular switch platform for rapid, sensitive, and point-of-care detection of SARS-CoV-2.
• To engineer a chimeric construct combining aequorin and an ACE2-derived peptide for specific viral target recognition.

Main Methods:

• Engineered a chimeric construct (CovSwitch) integrating aequorin and a peptide sequence from human ACE2.
• Utilized bioluminescence intensity to quantify viral load in a dose-dependent manner.
• Performed site-directed mutagenesis, immunoassays, and in silico analyses to elucidate the interaction mechanism.

Main Results:

• CovSwitch demonstrated high sensitivity for SARS-CoV-2 detection, with a linear range from Ct 19 to 37 (2.7 × 10^7 to 10 copies/μl) and a detection limit of Ct 40.54 (<10 copies/μL).
• The platform exhibited high selectivity and successfully detected SARS-CoV-2 in clinical nasal fluid samples.
• Specific interaction between CovSwitch and SARS-CoV-2 RBD was confirmed, leading to enhanced bioluminescence and optimized binding conformation.

Conclusions:

• Photoprotein-based switch systems show significant potential as highly sensitive, selective, and label-free platforms for SARS-CoV-2 detection.
• The developed single-step platform is adaptable for detecting other viral targets, offering broad diagnostic applications.