Rapid Detection of Anti-SARS-CoV-2 Antibody Using a Selenium Nanoparticle-Based Lateral Flow Immunoassay

Abstract

Coronavirus disease 2019 is an infectious disease caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). SARS-CoV-2 is highly transmissible. Early and rapid testing is necessary to effectively prevent and control the outbreak. Detection of SARS-CoV-2 antibodies with lateral flow immunoassay can achieve this goal. In this study, SARS-CoV-2 nucleoprotein (NP) was expressed and purified. We used the selenium nanoparticle as the labeling probe coupled with the NP to prepare an antibody (IgM and IgG) detection kit. The detection limit, cross reaction, sensitivity and specificity of the kit is verified. Separate detection of IgM and IgG, such as in this assay, was performed in order to reduce mutual interference and improve the accuracy of the test results.The final purity of NP was 91.83%. Selenium nanoparticle and NP successfully combined with stable effect. The LOD of the kit was 20 ng/mL for anti-NP IgG and 60 ng/mL for anti-NP IgM, respectively. The kit does not cross reaction with RF. The sensitivity of the kit was 94.74% and the specificity was 96.23%. The assay kit does not require any special device for reading the results and the readout is a simple color change that can be evaluated with the naked eye. This kit is suitable for rapid and real-time detection of the SARS-CoV-2 antibody IgG and IgM.

Fig. 1.

Diagram and components of the SARS-CoV-2 antibody immunoassay test strip (a) and visual assessment guidelines for interpreting the test strip results (b). Abbreviation: SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; C: Control line; G: Test line for IgG detection; M: Test line for IgM detection.

Fig. 2.

SARS-CoV-2 recombinant nucleoprotein purification results. (a) SDS-PAGE, (b) Purity verification byHPLC, Abbreviation: 1) Bacterial culture before induction. 2) Bacterial culture after induction. 3) Precipitate from centrifuged bacterial culture after induction. 4) Supernatant from centrifuged bacterial culture after induction. 5) Flow-through of nickel column supernatant. 6) Eluate of the target protein—nucleoprotein conjugate. 7) The nucleoprotein by ion exchange. SARS-CoV-2, severe acute respiratory syndrome coronavirus.

Fig. 3.

pH value determination for selenium nanoparticle-conjugated recombinant nucleoprotein. Photographs of centrifuge, resuspending (a) the test strip naked results (b), auxiliary judgment result (c), morphology of Se-NP (d), Se-NP-nucleoprotein (e), size analysis (f) and combined effect (g) are shown. C: Control line; G: Test line for IgG detection; M: Test line for IgM detection; PDI: Polymer dispersity index; Se-NP: Selenium nanoparticle.

Fig. 4.

Selection of conjugating and coating conditions. Detection result for IgG (a) and IgM (b) with fetal bovine serum; detection result for IgG (c) and IgM (d) with two health human serum; C line: Control line; G line: Test line for IgG detection; M line: Test line for IgM detection; C: Concentration.

Fig. 5.

Limit of detection of the kit for IgG and IgM detection. Result of detection for IgG (a) and IgM (c) by naked eyes; Analysis result of detection for IgG (b) and IgM (d) by GraphPad Prism 8. C: Control line; G: Test line for IgG detection; M: Test line for IgM detection.

Fig. 6.

Cross reaction results of anti-SARS-CoV-2 IgG (a) and IgM (b) for rheumatoid factor. SARS-CoV-2, severe acute respiratory syndrome coronavirus 2. P: Positive; N: Negative; C: Control line; G: Test line for IgG detection; M: Test line for IgM detection.

Fig. 7.

Clinical sample detection results of anti-SARS-CoV-2 IgG (a) and IgM (b) by naked eyes; Analysis result of detection for IgG (c) and IgM (d) by GraphPad Prism 8. SARS-CoV-2, severe acute respiratory syndrome coronavirus 2. C: Control line; G: Test line for IgG detection; M: Test line for IgM detection.