Bacterial ligands as flexible and sensitive detectors in rapid tests for antibodies to SARS-CoV-2

Simone Cavalera,Celeste Cagnazzo,Cristina Guiotto,Sergio Rosati,Marco Denina,Annagloria Palazzo,Fabio Di Nardo,Roberto Pisano,Laura Anfossi,Fiora Artusio,Franca Fagioli,Barbara Colitti,Claudio Baggiani,Matteo Chiarello,Thea Serra

doi:10.1007/s00216-022-03939-2

Abstract

Lateral flow immunoassay (LFIA) is widely employed as point-of-care tests (POCT) for the diagnosis of infectious diseases. The accuracy of LFIA largely depends on the quality of the immunoreagents used. Typical LFIAs to reveal the immune response to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) employ anti-human immunoglobulin (hIG) antibodies and recombinant viral antigens, which usually are unstable and poorly soluble. Broad selective bacterial proteins, such as Staphylococcal protein A (SpA) and Streptococcal protein G (SpG) can be considered alternatives to anti-hIG to increase versatility and sensitivity of serological LFIAs because of their high binding capacity, interspecies reactivity, and robustness. We developed two colorimetric LFA devices including SpA and SpG linked to gold nanoparticles (GNP) as detectors and explored the use of a specific, stable, and soluble immunodominant fraction of the nucleocapsid protein from SARS-CoV-2 as the capturing agent. The optimal amount of SpA-GNP and SpG-GNP conjugates and the protein-to-GNP ratios were defined through a full factorial experimental design to maximize the diagnostic sensitivity of the LFIAs. The new LFA devices were applied to analyze 105 human serum samples (69 positive and 36 negatives according to reference molecular diagnostic methods). The results showed higher sensitivity (89.9%, 95% CI 82.7–97.0) and selectivity (91.7%, 82.6–100) for the SpA-based compared to the SpG-based LFA. In addition, 18 serum samples from cats and dogs living with COVID-19 patients were analyzed and 14 showed detectable levels of anti-SARS-CoV-2 antibodies, thus illustrating the flexibility of the SpA- and SpG-based LFAs.Graphical abstract

Highlights

After 2 years from the outbreak of the novel coronavirus, SARS-CoV-2 pandemic, the situation is still on the razor-edge
We propose two LFA devices able to detect the immune response to SARS-CoV-2 in humans and pets by using Staphylococcal protein A (SpA) and Streptococcal protein G (SpG) labelled with gold nanoparticles (GNPs) as the probes
The two antibody-LFAs proposed in this work were both able to diagnose the immune response to SARS-CoV-2 in humans and companion animals

Summary

Introduction

After 2 years from the outbreak of the novel coronavirus, SARS-CoV-2 pandemic, the situation is still on the razor-edge. It is worth noting that the trend to discriminate between anti-SARS-CoV-2 immunoglobulin M (IgM) and immunoglobulin G (IgG), typical of the spring 2020 rapid serological test generation, has been largely explored but it is still under discussion. The effects of using Staphylococcal protein A (SpA) and Streptococcal protein G (SpG) as detection [14, 15] or capturing elements [16] on antibody testing have been explored Another interesting benefit of bacterial bioligands is their broad selectivity towards immunoglobulins of different animal species. The use of SpG and SpA as detection ligands provides the required flexibility to apply the same LFA device to detect the serological response in different animal species Such bioligands are used for purification of immunoglobulins [17]. The use of SpA and SpG as probes in LFA has been reported in several scientific works and for commercially available devices [15, 16, 21]

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Results

Conclusion