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用于快速检测灭活传染性脾肾坏死病毒疫苗中MCP抗原浓度的双抗体夹心酶联免疫吸附测定法的开发

Development of a Double-Antibody Sandwich ELISA for Rapid Detection of the MCP Antigen Concentration in Inactivated ISKNV Vaccines.

作者信息

Liang Hongru, Zhang Lixi, Fu Xiaozhe, Lin Qiang, Liu Lihui, Niu Yinjie, Luo Xia, Huang Zhibin, Li Ningqiu

机构信息

Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture and Rural Affairs, Guangdong Province Key Laboratory of Aquatic Animal Immune Technology, Guangzhou 510380, China.

出版信息

Vaccines (Basel). 2021 Nov 2;9(11):1264. doi: 10.3390/vaccines9111264.

DOI:10.3390/vaccines9111264
PMID:34835196
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8623861/
Abstract

Infectious spleen and kidney necrosis virus (ISKNV) resulted in severe systemic diseases with high morbidity and mortality in Siniperca chuatsi. Vaccination is the primary method for effective prevention and control of these diseases. The development of inactivated ISKNV vaccines made some progress, but the technique of quality evaluation is scarce. Herein, a measurement of the MCP (major capsid protein) antigen concentration for the inactivated ISKNV vaccine was developed by double-antibody sandwich ELISA. Firstly, mouse monoclonal antibodies against ISKNV particles and MCP were generated. Then, a double-antibody sandwich ELISA was developed using the monoclonal antibody 1C8 1B9 as the capture antibody and Biotin-3B12 6B3 as the detection antibody. A standard curve was generated using the MCP concentration versus OD value with the linear range of concentration of 4.69~300 ng/mL. The assay sensitivity was 0.9 ng/mL. The antigen content of three batches of inactivated ISKNV vaccines was quantitatively detected using the double-antibody sandwich ELISA. The results showed that MCP antigen contents of inactivated ISKNV vaccines were positively correlated with the viral titers. The newly established double-antibody sandwich ELISA provided a useful tool for the detection of antigen quality for ISKNV inactivated vaccines.

摘要

传染性脾肾坏死病毒（ISKNV）可导致鳜鱼出现严重的全身性疾病，发病率和死亡率都很高。疫苗接种是有效预防和控制这些疾病的主要方法。灭活ISKNV疫苗的研发取得了一些进展，但质量评估技术却很匮乏。在此，通过双抗体夹心ELISA法建立了一种针对灭活ISKNV疫苗的主要衣壳蛋白（MCP）抗原浓度的检测方法。首先，制备了针对ISKNV病毒颗粒和MCP的小鼠单克隆抗体。然后，以单克隆抗体1C8 1B9作为捕获抗体，生物素化的3B12 6B3作为检测抗体，建立了双抗体夹心ELISA法。以MCP浓度对OD值绘制标准曲线，浓度线性范围为4.69~300 ng/mL。该检测方法的灵敏度为0.9 ng/mL。利用双抗体夹心ELISA法定量检测了三批灭活ISKNV疫苗的抗原含量。结果表明，灭活ISKNV疫苗的MCP抗原含量与病毒滴度呈正相关。新建立的双抗体夹心ELISA法为检测ISKNV灭活疫苗的抗原质量提供了一种有用的工具。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/3f9485288ad1/vaccines-09-01264-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/dc19653770f3/vaccines-09-01264-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/f19e7a20f8d2/vaccines-09-01264-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/4e765ed5c693/vaccines-09-01264-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/032ff8c45a85/vaccines-09-01264-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/b864b817ea11/vaccines-09-01264-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/3f9485288ad1/vaccines-09-01264-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/dc19653770f3/vaccines-09-01264-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/f19e7a20f8d2/vaccines-09-01264-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/4e765ed5c693/vaccines-09-01264-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/032ff8c45a85/vaccines-09-01264-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/b864b817ea11/vaccines-09-01264-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cd16/8623861/3f9485288ad1/vaccines-09-01264-g006.jpg

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