3D Film-Like Nanozyme for Ultrasensitive Detection of Respiratory Viruses

Respiratory viral infections pose a significant challenge to global public health, and rapid and accurate detection is crucial for timely diagnosis and treatment. Traditional detection methods such as ELISA and PCR have limitations including complex operations and long processing times. Lateral flow immunoassay (LFA) test strips have become an ideal instant detection technology due to their simplicity and speed, but sensitivity needs improvement. The introduction of nanozymes provides a new approach to enhance LFA performance, but designing efficient nanozymes and integrating them into LFA for ultra-sensitive detection remains a challenge. Therefore, developing novel high-efficiency nanozymes and applying them to LFA for ultra-sensitive rapid detection of respiratory viruses is of great scientific significance and application value for improving disease diagnosis efficiency and public health.

Recently, Professor Gu Bing and his research team at Guangdong Provincial People’s Hospital developed a 3D film-like nanozyme (GO−Pt30−AuPt5) for ultra-sensitive immunochromatographic detection of respiratory viruses. This nanozyme significantly improves detection sensitivity by assembling platinum and gold@platinum nanoparticles on a two-dimensional graphene oxide nanosheet. Studies show that GO−Pt30−AuPt5 can achieve simultaneous detection of two important respiratory viruses with a sensitivity of 1 pg/mL, about 100 times higher than traditional methods. Clinical validation demonstrated excellent performance in 49 real samples, showcasing its broad application potential in pathogen screening. The related results were published in ACS Nano under the title “3D Film-Like Nanozyme with a Synergistic Amplification Effect for the Ultrasensitive Immunochromatographic Detection of Respiratory Viruses” (CAS Top Journal).

GO−Pt30−AuPt5 nanozyme is prepared by assembling 30 nm platinum nanoparticles and 5 nm gold@platinum nanoparticles on a two-dimensional graphene oxide nanosheet. This nanozyme exhibits a three-dimensional structure with good flexibility and high surface area, allowing for high dispersion in water. Research shows that GO−Pt30−AuPt5 exhibits strong chromogenic and catalytic activity in catalytic reactions, effectively enhancing the detection sensitivity for respiratory viruses, demonstrating its potential for rapid, simple, ultra-sensitive pathogen screening in clinical samples.

Figure 1. Preparation and characterization of film-like GO-Pt30-AuPt5 nanozyme.

This study developed various types of nanozymes to evaluate their enzymatic activity, including GO−Pt30, GO−AuPt5, GO−Pt30−Pt30, GO−AuPt5−AuPt5, and GO−Pt30−AuPt5. Results indicate that GO−Pt30−AuPt5 nanozyme exhibits the highest catalytic activity, significantly outperforming other types. The differences in catalytic performance among different nanozymes mainly stem from their structure and composition. By combining 30 nm platinum nanoparticles with 5 nm gold@platinum nanoparticles, GO−Pt30−AuPt5 not only increases catalytic sites and reaction interfaces but also significantly enhances its catalytic efficiency, exhibiting high sensitivity and excellent catalytic capability. Additionally, the catalytic efficiency of GO−Pt30−AuPt5 far exceeds that of traditional peroxidase, demonstrating its broad application potential in biosensing and disease detection.

Figure 2. Evaluation of peroxidase-like activity of GO-based film nanozymes.

GO-Pt30-AuPt5 nanozyme is prepared through PEI-mediated self-assembly, combining the excellent catalytic performance of platinum and gold@platinum nanoparticles. This nanozyme achieves targeted binding through antibody modification, constructing a double-antibody sandwich immunodetection model to enhance specificity and signal amplification for SARS-CoV-2 and H1N1.

Figure 3. Schematic diagram of the preparation of antibody-modified GO-Pt30-AuPt5, and the structural design and detection model based on this nanozyme for detecting SARS-CoV-2 and H1N1.

GO-Pt30-AuPt5 nanozyme exhibits excellent catalytic activity on the AEC substrate, rapidly decomposing H2O2 to generate OH radicals, thereby catalyzing the oxidation reaction of AEC to form an insoluble brown-red precipitate. This catalytic process not only significantly enhances the detection signal but also improves the sensitivity and specificity for the target viruses, aiming to achieve rapid and sensitive detection of SARS-CoV-2 and H1N1.

Figure 4. Catalytic activity characterization of GO-Pt30-AuPt5 nanozyme on LFA strips.

When detecting different concentrations of SARS-CoV-2 and H1N1, GO-Pt30-AuPt5 nanozyme demonstrates excellent performance. Prior to the catalytic reaction, the nanozyme produces a significant black signal at high virus concentrations, which gradually weakens as the virus concentration decreases, ultimately becoming colorless at concentrations below 0.5 ng/mL. After the catalytic reaction, the signal intensity of the T line significantly increases, greatly expanding the detection range. In catalytic mode, the detection limits of this method are 0.0013 ng/mL and 0.00084 ng/mL, showing extremely high sensitivity and a wide dynamic detection range, suitable for rapid and accurate detection of respiratory viruses.

Figure 5. Detection effect of GO–Pt30–AuPt5-LFA for SARS-CoV-2 and H1N1.

In the detection of real clinical samples, GO-Pt30-AuPt5 nanozyme successfully identified positive pharyngeal swab samples for H1N1 and SARS-CoV-2, with all positive samples showing significant T line signals, indicating a 100% detection rate, validating its accuracy and reliability in clinical applications.

Figure 6. Detection results of GO–Pt30–AuPt5-LFA for two target viruses in actual samples.

In summary, this study successfully developed the GO-Pt30-AuPt5 three-dimensional nanozyme, significantly enhancing the detection sensitivity and range for H1N1 and SARS-CoV-2. The effective application of this nanozyme in real clinical samples demonstrates its potential for rapid and accurate virus detection, providing reliable technical support for pathogen screening.