RESUMEN
Rapid and sensitive antigen detection using a lateral flow immunoassay (LFIA) is crucial for diagnosing infectious diseases due to its simplicity, speed, and user-friendly features. However, it remains a critical issue to explore specific biorecognition elements and powerful signal amplification. In this study, taking SARS-CoV-2 as a proof of concept, a specific peptide, WFLNDSELIML, binding to the SARS-CoV-2 spike (S) antigen was identified by a nonamplified biopanning method, which exhibited high affinity to the target, with a dissociation constant of 9.29 ± 1.55 nM. Molecular docking analysis reveals that this peptide binds to the N-terminal domain of the SARS-CoV-2 S antigen. Then, using this peptide as a capture probe and angiotensin-converting enzyme 2 as a detection probe, a peptide-based lateral flow immunoassay (pLFIA) for the sensitive detection of the SARS-CoV-2 S antigen without any antibody was developed, for which a polydopamine nanosphere (PDA)@MnO2 nanocomposite with excellent oxidase-like activity was used as a colorimetric label, exhibiting dual-mode remarkable signal amplification of natural melanin and on-demand nanozyme catalytic enhancement. The PDA@MnO2-based pLFIA is capable of detecting the SARS-CoV-2 S antigen with a limit of detection of 8.01 pg/mL, which is 18.7 times lower than that of a conventional pLFIA tagged with gold nanoparticles. Additionally, the as-proposed PDA@MnO2-based pLFIA can detect up to 150 transduction units/mL SARS-CoV-2 pseudoviruses spiked in saliva samples. Given the outstanding analytical performance, the proposed PDA@MnO2-based pLFIA may offer a reliable option for the rapid diagnosis of SARS-CoV-2.
Asunto(s)
Antígenos de Grupos Sanguíneos , COVID-19 , Nanopartículas del Metal , Humanos , COVID-19/diagnóstico , Oro , Compuestos de Manganeso , Simulación del Acoplamiento Molecular , Óxidos , SARS-CoV-2 , InmunoensayoRESUMEN
The COVID-19 pandemic has led to a global crisis with devastating effects on public healthcare and the economy. Sensitive detection of SARS-CoV-2 is the key to diagnose and control its spread. The spike (S) protein is an abundant viral transmembrane protein and a suitable target protein for the selective recognition of SARS-CoV-2. Here, we report that with bovine serum albumin prescreening, a specific phage peptide targeting SARS-CoV-2 S1 protein was biopanned with the pIII phage display library. The identified phage #2 expressing the peptide (amino acid sequence: NFWISPKLAFAL) shows high affinity to the target with a dissociation constant of 3.45 ± 0.58 nM. Furthermore, the identified peptide shows good specificity with a binding site at the N-terminal domain of the S1 subunit through a hydrogen bond network and hydrophobic interaction, supported by molecular docking. Then, a sandwiched phage-based enzyme-linked chemiluminescence immunoassay (ELCLIA) was established by using phage #2 as a bifunctional probe capable of SARS-CoV-2 S1 antigen recognition and signal amplification. After optimizing the conditions, the proposed phage ELCLIA exhibited good sensitivity, and as low as 78 pg/mL SARS-CoV-2 S1 could be detected. This method can be applied to detect as low as 60 transducing units (TU)/mL SARS-CoV-2 pseudovirus in 50% saliva. Therefore, specific phage peptides have good prospects as powerful biological recognition probes for immunoassay detection and biomedical applications.
Asunto(s)
Bacteriófagos , COVID-19 , COVID-19/diagnóstico , Humanos , Inmunoensayo , Luminiscencia , Simulación del Acoplamiento Molecular , Pandemias , Péptidos , SARS-CoV-2RESUMEN
Gold nanoplates (AuNPTs) exhibit outstanding photothermal conversion efficiency (68.5%) and peroxidase-like activity. The combination of the very low H2O2 concentration (0.1 mM) and the low AuNPT dosage (50 µg mL-1) with 808 nm laser irradiation (1 W cm-2, 3 min) shows excellent synergistic antibacterial ability and healing of MRSA-infected wounds in vivo.