Colorimetric and Raman dual-mode lateral flow immunoassay detection of SARS-CoV-2 N protein antibody based on Ag nanoparticles with ultrathin Au shell assembled onto Fe3O4 nanoparticles

Serological antibody tests are useful complements of nuclei acid detection for SARS-CoV-2 diagnosis, which can significantly improve diagnostic accuracy. However, antibody detection in serum or plasma remains challenging to do with high sensitivity. In this study, Ag nanoparticles with ultra-thin Au shells embedded with 4-mercaptobenzoic acid (MBA) (Ag-MBA@Au) were manufactured and then assembled onto Fe3O4 surface by electrostatic interaction to construct the Fe3O4-Ag-M(BA)@Au nanoparticles (NPs) with magnetic-Raman-colorimetric properties. Based on the composite nanoparticles, a colorimetric and Raman dual-mode lateral flow immunoassay (LFIA) for ultrasensitive identification of SARS-CoV-2 nucleocapsid (N) protein antibody was constructed. The magnetic nanoparticles (Fe3O4 NPs) were acted as the core and coated a layer of Ag-M(BA)@Au particles on the surface by electrostatic interaction to prepare Fe3O4-Ag-M(BA)@Au NPs, which can amplify the SERS signal due to multiple Ag-M(BA)@Au particles concentrated on a single magnetic nanoparticle. Moreover, the Fe3O4-Ag-M(BA)@Au NPs facilitated pre-purifying sample using magnetic separation, and complex matrix interference would be greatly decreased in the detection. The Fe3O4-Ag-M(BA)@Au NPs modified with N protein recognized and bound with N protein antibodies, which were trapped on the T-line, forming color band for observing detection. Under optimal conditions, the N protein antibodies could be qualitatively detected in colorimetric mode with the visual limit of 10(-8) mg/mL and quantitatively detected by SERS signals between 10(-6) and 10(-)(10) mg /mL with 0.08 pg/mL detection limit. The coefficients variations (CV) of intra-assay was 8.0%, whereas of inter-assay was 11.7%, confirming of good reproducibility. Finally, this approach was able to discriminate between positive, negative, and weakly positive samples when detecting 107 clinical serum samples. The process enables highly sensitive quantitative assays that are valuable for evaluating disease processes and guiding treatment.

Automated summary

In this study, a novel magnetic-Raman-colorimetric nanoparticle was developed for highly sensitive detection of SARS-CoV-2 nucleocapsid protein antibodies. The method exhibited high accuracy, sensitivity, and reproducibility, and could differentiate different serum samples. This approach is valuable for evaluating disease processes and guiding treatment.