Development of a biosafe ELISA-based platform for assessing immunogenicity in the production of an inactivated whole-virion coronavirus vaccine

Abstract

BACKGROUND: SARS-CoV-2 vaccine immunogenicity is evaluated in neutralization test with live virus. It is performed in a biosafety level 3 zone because requires live virus stage. Therefore, control laboratories should be certified for this class of work. The development of technology based on enzyme-linked immunosorbent assay as an analogue of the neutralization reaction makes it possible to create an immunobiological product in a shorter time and in conditions without special requirements for control laboratories.

AIM: Development of an enzyme-linked immunosorbent assay for assessing SARS-CoV-2 vaccine immunogenicity by measuring neutralizing antibodies production in immunized animals.

MATERIALS AND METHODS: Recombinant receptor-binding domain fused to a С-terminal hexahistidine sequence was produced in Escherichia coli cells and purified via metal-affinity chromatography on WorkBeads NiMAC (Bio-Works). Purified protein was used in enzyme-linked immunosorbent assay as an antigen for sorption. The sera of mice immunized with the vaccine preparation were tested for neutralizing activity against the SARS-CoV-2, as well as in the developed enzyme-linked immunosorbent assay.

RESULTS: Sera with high neutralizing titers showed a high degree of binding to recombinant receptor-binding domain fused to a С-terminal hexahistidine sequence in enzyme-linked immunosorbent assay, while sera from non-immunized animals or sera with neutralization titers less than 1:8 were not reactive in enzyme-linked immunosorbent assay. The Spearman and Pearson correlation coefficients for neutralization test titers and optical density in enzyme-linked immunosorbent assay were 0.759 and 0.76, respectively. The developed assay can be used as a semi-quantitative method for assessing the immunogenicity of a vaccine against coronavirus infection.

CONCLUSIONS: The developed platform makes it possible to reliably assess the immunogenicity of an inactivated coronavirus vaccine under conditions that do not require a high biosafety conditions.

Keywords

SARS-CoV2 / vaccines / immunogenicity / neutralization / coronavirus / biosafety

References

[1] Talic S, Shah S, Wild H, et al. Effectiveness of public health measures in reducing the incidence of COVID-19, SARS-CoV-2 transmission, and COVID-19 mortality: systematic review and meta-analysis. BMJ. 2021;375:e068302. DOI: 10.1136/bmj-2021-068302

[2] Talic S., Shah S., Wild H. et al. Effectiveness of public health measures in reducing the incidence of COVID-19, SARS-CoV-2 transmission, and COVID-19 mortality: systematic review and meta-analysis // BMJ. 2021. Vol. 375. P. e068302. DOI: 10.1136/bmj-2021-068302

[3] Kozlovskaya LI, Piniaeva AN, Ignatyev GM, et al. Long-term humoral immunogenicity, safety and protective efficacy of inactivated vaccine against COVID-19 (CoviVac) in preclinical studies. Emerg Microbes Infect. 2021;10(1):1790–1806. DOI: 10.1080/22221751.2021.1971569

[4] Kozlovskaya L.I., Piniaeva A.N., Ignatyev G.M. et al. Long-term humoral immunogenicity, safety and protective efficacy of inactivated vaccine against COVID-19 (CoviVac) in preclinical studies // Emerg. Microbes Infect. 2021. Vol. 10, No. 1. P. 1790–1806. DOI: 10.1080/22221751.2021.1971569

[5] Joseph T, Phyu S, Se-Thoe SY, Chu JJH. Biorisk management for SARS-CoV-2 research in a biosafety level-3 core facility. Methods Mol Biol. 2022;2452:441–464. DOI: 10.1007/978-1-0716-2111-0_24

[6] Joseph T., Phyu S., Se-Thoe S.Y., Chu J.J.H. Biorisk management for SARS-CoV-2 research in a biosafety level-3 core facility // Methods Mol. Biol. 2022. Vol. 2452. P. 441–464. DOI: 10.1007/978-1-0716-2111-0_24

[7] Kim DK, Kim HY, Kim JY, et al. Development of an in vitro antigen-detection test as an alternative method to the in vivo plaque reduction neutralization test for the quality control of Japanese encephalitis virus vaccine. Microbiol Immunol. 2012;56(7):463–471. DOI: 10.1111/j.1348-0421.2012.00462.x

[8] Kim D.K., Kim H.Y., Kim J.Y. et al. Development of an in vitro antigen-detection test as an alternative method to the in vivo plaque reduction neutralization test for the quality control of Japanese encephalitis virus vaccine // Microbiol. Immunol. 2012. Vol. 56, No. 7. P. 463–471. DOI: 10.1111/j.1348-0421.2012.00462.x

[9] Balingit JC, Phu Ly MH, Matsuda M, et al. A simple and high-throughput ELISA-based neutralization assay for the determination of anti-flavivirus neutralizing antibodies. Vaccines (Basel). 2020;8(2):297. DOI: 10.3390/vaccines8020297

[10] Balingit J.C., Phu Ly M.H., Matsuda M. et al. A simple and high-throughput ELISA-based neutralization assay for the determination of anti-flavivirus neutralizing antibodies // Vaccines (Basel). 2020. Vol. 8, No. 2. P. 297. DOI: 10.3390/vaccines8020297

[11] Jelektroforez v poliakrilamidnom gele. OFS.1.2.3.0023.15. Gosudarstvennaya farmakopeya Rossiiskoi Federatsii. XIV. Vol. I. (In Russ.)

[12] Электрофорез в полиакриламидном геле. ОФС.1.2.3.0023.15 // Государственная фармакопея Российской Федерации XIV. Т. I.

RIGHTS & PERMISSIONS

Eco-Vector