A fluorescent microspheres-based microfluidic test system for the detection of immunoglobulin G to SARS-CoV-2

Ruslan I. Shakurov , Yaroslav D. Shansky , Kirill A. Prusakov , Svetlana V. Sizova , Stepan P. Dudik , Lyudmila V. Plotnikova , Valentin A. Manuvera , Dmitry V. Klinov , Vassili N. Lazarev , Julia A. Bespyatykh , Dmitriy V. Basmanov

Journal of Clinical Practice ›› 2023, Vol. 14 ›› Issue (1) : 44 -53.

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Journal of Clinical Practice ›› 2023, Vol. 14 ›› Issue (1) : 44 -53. DOI: 10.17816/clinpract278280
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A fluorescent microspheres-based microfluidic test system for the detection of immunoglobulin G to SARS-CoV-2

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Abstract

Background: The pandemic of the new coronavirus infection, COVID-19, is currently ongoing in the world. Over the years, the pathogen, SARS-CoV-2, has undergone a series of mutational genome changes, which has led to the spread of various genetic variants of the virus. Meanwhile, the methods used to diagnose SARS-CoV-2, to establish the disease stage and to assess the immunity, are nonspecific to SARS-CoV-2 variants and time-consumable. Thus, the development of new methods for diagnosing COVID-19, as well as their implementation in practice, is currently an important direction. In particular, application of systems based on chemically modified fluorescent microspheres (with a multiplex assay for target protein molecules) opens great opportunities.

Aim: development of a microfluidic diagnostic test system based on fluorescent microspheres for the specific detection of immunoglobulins G (IgG) to SARS-CoV-2.

Methods: A collection of human serum samples was characterized using enzyme-linked immunosorbent assay (ELISA) and commercially available reagent kits. IgG to SARS-CoV-2 in the human serum were detected by the developed immunofluorescent method using microspheres containing the chemically immobilized RBD fragment of the SARS-CoV-2 (“Kappa” variant) viral S-protein.

Results: The level of IgG in the blood serum of recovered volunteers was 9-300 times higher than that in apparently healthy volunteers, according to ELISA (p<0.001). Conjugates of fluorescent microspheres with the RBD-fragment of the S-protein, capable of specifically binding IgG from the blood serum, have been obtained. The immune complexes formation was confirmed by the fluorescence microscopy data; the fluorescence intensity of secondary antibodies in the immune complexes formed on the surface of microspheres was proportional to the content of IgG (r 0.963). The test system had a good predictive value (AUC 70.3%).

Conclusion: A test system has been developed, based on fluorescent microspheres containing the immobilized RBD fragment of the SARS-CoV-2 S-protein, for the immunofluorescent detection of IgG in the human blood serum. When testing the system on samples with different levels of IgG to SARS-CoV-2, its prognostic value was shown. The obtained results allow us to present the test system as a method to assess the level of immunoglobulins to SARS-CoV-2 in the human blood serum for the implementation in clinical practice. The test system can also be integrated into various microfluidic systems to create chips and devices for the point-of-care diagnostics.

Keywords

enzyme-linked immunosorbent assay / ELISA / COVID-19 testing / personalized medicine

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Ruslan I. Shakurov, Yaroslav D. Shansky, Kirill A. Prusakov, Svetlana V. Sizova, Stepan P. Dudik, Lyudmila V. Plotnikova, Valentin A. Manuvera, Dmitry V. Klinov, Vassili N. Lazarev, Julia A. Bespyatykh, Dmitriy V. Basmanov. A fluorescent microspheres-based microfluidic test system for the detection of immunoglobulin G to SARS-CoV-2. Journal of Clinical Practice, 2023, 14(1): 44-53 DOI:10.17816/clinpract278280

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Funding

Научное исследование проведено при поддержке гранта Президента Российской ФедерацииThis work was supported by the grant of the President of Russian Federation(МК-2988.2022.3)

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Shakurov R.I., Shansky Y.D., Prusakov K.A., Sizova S.V., Dudik S.P., Plotnikova L.V., Manuvera V.A., Klinov D.V., Lazarev V.N., Bespyatykh J.A., Basmanov D.V.

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