A uniform quantitative enzyme-linked immunosorbent assay for Coxsackievirus A16 antigen in vaccine

ABSTRACT

Coxsackievirus A16 (CV-A16), one of major etiological agents of hand, foot and mouth disease (HFMD), causes outbreaks of the disease in young children all over the world. In order to promote the prevention and control of HFMD, the research and development of CV-A16 vaccine have been carried out in China. However, due to lacking of a recognized CV-A16 antigen detection method, the evaluation and quality control (QC) of vaccine effectiveness are greatly limited. In this study, we established a quantitative enzyme-linked immunosorbent assay (Q-ELISA) to determine the antigen concentration in CV-A16 vaccines that can be applied in manufacturing in China. A neutralizing antibody 16E1 was used as a capture antibody that can bind to various CV-A16 antigens of different subgenotypes, and an antiserum from CV-A16-immunized rabbit conjugated by HRP was suitable for detecting and quantifying CV-A16 antigens. The Q-ELISA was validated for specificity, linearity, accuracy, precision and robustness by using the CV-A16 antigen national standard (NS). Furthermore, we utilized the Q-ELISA to quantify antigen contents of vaccine bulks from six manufacturers and other intermediate products from one manufacturer. The results indicated that the Q-ELISA can satisfy the requirements of QC for all manufacturers involved.

KEYWORDS:

• Coxsackievirus A16 (CV-A16)
• antigen
• Q-ELISA
• neutralizing antibody
• vaccine

Acknowledgments

Manufacturers devoted to developing CV-A16 inactivated vaccine were involved in this applicability validation of this study, including WuHan Institute of Biological Products Co. Ltd, Wuhan, PR China; National Vaccine and Serum Institute, Beijing, PR China; Chinese Academy of Medical Sciences, Kunming, PR China; Aimei Convac BioPharm (Jiangsu) Co. Ltd, Taizhou, PR China; and Beijing Zhifei Lvzhu Biopharmaceutical Co. Ltd., Beijing, PR China.

Additional information

Funding

This work was supported by a grant from the National Science and Technology Major Projects for Major New Drugs Innovation and Development (2018ZX09711003-005-003), the National Natural Science Foundation of China (31670933). The funders had no role in the study design, data collection and analysis, decision to publish or preparation of the manuscript.