Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

Development and Validation of an Enzyme-Linked Immunosorbent Assay-Based Protocol for Evaluation of Respiratory Syncytial Virus Vaccines

Eliel Nham
,
A-Yeung Jang
ORCID logo ,
Hyun Jung Ji
,
Ki Bum Ahn
,
Joon-Yong Bae
,
Man-Seong Park
,
Hakjun Hyun
,
Jin Gu Yoon
ORCID logo ,
Hye Seong
,
Ji Yun Noh
,
Hee Jin Cheong
,
Woo Joo Kim
ORCID logo ,
Ho Seong Seo
* ORCID logo ,
Joon Young Song
* ORCID logo
Version 1 : Received: 22 June 2023 / Approved: 23 June 2023 / Online: 23 June 2023 (08:36:32 CEST)

How to cite: Nham, E.; Jang, A.; Ji, H.J.; Ahn, K.B.; Bae, J.; Park, M.; Hyun, H.; Yoon, J.G.; Seong, H.; Noh, J.Y.; Cheong, H.J.; Kim, W.J.; Seo, H.S.; Song, J.Y. Development and Validation of an Enzyme-Linked Immunosorbent Assay-Based Protocol for Evaluation of Respiratory Syncytial Virus Vaccines. Preprints 2023, 2023061646. https://doi.org/10.20944/preprints202306.1646.v1 Nham, E.; Jang, A.; Ji, H.J.; Ahn, K.B.; Bae, J.; Park, M.; Hyun, H.; Yoon, J.G.; Seong, H.; Noh, J.Y.; Cheong, H.J.; Kim, W.J.; Seo, H.S.; Song, J.Y. Development and Validation of an Enzyme-Linked Immunosorbent Assay-Based Protocol for Evaluation of Respiratory Syncytial Virus Vaccines. Preprints 2023, 2023061646. https://doi.org/10.20944/preprints202306.1646.v1

Abstract

Recently, two respiratory syncytial virus (RSV) vaccines based on the prefusion F (pre-F) antigen were approved in the United States. We developed and validated an enzyme-linked immunosorbent assay (ELISA)-based protocol for the practical and large-scale evaluation of RSV vaccines. Two modified pre-F proteins (DS-Cav1 and SC-TM) were produced by genetic recombination and replication using an adenoviral vector. The protocol was established by optimizing the concentrations of the coating antigen (pre-F proteins), secondary antibodies, and choice of blocking buffer. To validate the protocol, we examined its accuracy, precision, and specificity using serum samples from 150 participants across various age groups and the standard serum provided by the National Institute of Health. In the linear correlation analysis, coating concentrations of 5 and 2.5 μg/mL of DS-Cav1 and SC-TM, respectively, showed high coefficients of determination (r2 = 0.90). Concentrations of secondary antibodies (alkaline phosphatase-conjugated anti-human immunoglobulin G, diluted 1:2000) and blocking reagents (5% skim milk/PBS-T) were optimized to minimize non-specific reactions. High accuracy was observed for DS-Cav1 (r2 = 0.90) and SC-TM (r2 = 0.86). Further, both antigens showed high precision (coefficient of variation < 15%). Inhibition ELISA revealed cross-reactivity of antibodies against DS-Cav1 and SC-TM but not with the attachment protein.

Keywords

respiratory syncytial virus; RSV; ELISA; Vaccine; Immunogenicity

Subject

Medicine and Pharmacology, Epidemiology and Infectious Diseases

Copyright: This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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