IBDV单克隆抗体的制备及双抗体夹心ELISA检测方法的建立

doi:10.11843/j.issn.0366-6964.2025.07.035

畜牧兽医学报 ›› 2025, Vol. 56 ›› Issue (7): 3433-3441.doi: 10.11843/j.issn.0366-6964.2025.07.035

IBDV单克隆抗体的制备及双抗体夹心ELISA检测方法的建立

姜艳平¹^,²(), 刘薇², 宫浩阳², 蔡李萌², 李佳璇²^,³, 崔文²^,³, 周晗²^,³, 韩建春¹^,*(), 唐丽杰²^,³^,*()

1. 黑龙江省绿色食品科学研究院，哈尔滨 150028
2. 东北农业大学动物医学学院，哈尔滨 150030
3. 黑龙江省动物疾病防控与制剂创制重点实验室，哈尔滨 150030

收稿日期:2024-09-30 出版日期:2025-07-23 发布日期:2025-07-25
通讯作者: 韩建春,唐丽杰 E-mail:jiangyanping8198@163.com;hanjianchun@hotmail.com;tanglijie@163.com
作者简介:姜艳平(1981-)，女，黑龙江肇东人，高级实验师，博士，主要从事兽医微生物学的教学与研究，E-mailjiangyanping8198@163.com
基金资助:
“十四五”重点研发项目子课题(2022YFD1800304-05)

Preparation of Monoclonal Antibodies to IBDV and Establishment of a Double Antibody Sandwich ELISA Method for Detection

JIANG Yanping¹^,²(), LIU Wei², GONG Haoyang², CAI Limeng², LI Jiaxuan²^,³, CUI Wen²^,³, ZHOU Han²^,³, HAN Jianchun¹^,*(), TANG Lijie²^,³^,*()

1. Heilongjiang Green Food Science Research Institute, Harbin 150028, China
2. College of Veterinary Medicine, Northeast Agricultural University, Harbin 150030, China
3. Heilongjiang Key Laboratory for Animal Disease Control and Pharmaceutical Development, Harbin 150030, China

Received:2024-09-30 Online:2025-07-23 Published:2025-07-25
Contact: HAN Jianchun, TANG Lijie E-mail:jiangyanping8198@163.com;hanjianchun@hotmail.com;tanglijie@163.com

摘要/Abstract

摘要：

旨在建立一种快速有效的检测鸡传染性法氏囊病病毒(IBDV)的方法，本研究以IBDV全病毒作为免疫原制备单克隆抗体，以获得的单抗6G3作为捕获抗体，单抗4C12作为检测抗体，建立检测IBDV的双抗体夹心ELISA方法。结果表明，该ELISA方法具有良好的特异性，与REV、AEV、ALV和EDSV均无交叉反应；能检测病毒的最小量为1.585×10³ ELD₅₀；批间和批内重复性试验显示，该方法具有良好的重复性；通过该方法对49份临床样品进行检测，与商品化的IBDV抗原检测卡进行比较，符合率达94.18%。综上，本研究建立的双抗体夹心ELISA方法，具有良好的特异性、敏感性和重复性，可用于IBDV的快速检测，为制备商品化的IBDV检测试剂盒奠定基础。

关键词: 传染性法氏囊病毒, 单克隆抗体, 检测方法, 双抗体夹心ELISA

Abstract:

The aim of this study was to establish a rapid and effective method for detecting infections bursal disease virus (IBDV), monoclonal antibodies were prepared using the whole IBDV as the immunogen. Double antibody sandwich ELISA method was established using monoclonal antibody 6G3 as the capture antibody and monoclonal antibody 4C12 as the detection antibody for the detection of IBDV. The results showed that the ELISA method had good specificity and no cross-reaction with REV, AEV, ALV or EDSV. The sensitivity test showed that the minimum detectable amount of IBDV is 1.585×10³ELD₅₀. The results of inter and intra plate repeatability tests showed that this method had good repeatability. Forty-nine clinical samples were detected by the double antibody sandwich ELISA method and commercial IBDV antigen detection card, and the coincidence rate was 94.18%. In conclusion, the double-antibody sandwich ELISA method established in this study has good specificity, sensitivity and repeatability, and can be used for rapid detection of IBDV. Thus, laysing the foundation for the development of a commercial IBDV detection kit.

Key words: IBDV, monoclonal antibody, detection method, double antibody sandwich ELISA

中图分类号:

S852.65

姜艳平, 刘薇, 宫浩阳, 蔡李萌, 李佳璇, 崔文, 周晗, 韩建春, 唐丽杰. IBDV单克隆抗体的制备及双抗体夹心ELISA检测方法的建立[J]. 畜牧兽医学报, 2025, 56(7): 3433-3441.

JIANG Yanping, LIU Wei, GONG Haoyang, CAI Limeng, LI Jiaxuan, CUI Wen, ZHOU Han, HAN Jianchun, TANG Lijie. Preparation of Monoclonal Antibodies to IBDV and Establishment of a Double Antibody Sandwich ELISA Method for Detection[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(7): 3433-3441.

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亚型Subtype	单抗Monoclonal antibody
IgG1κ	3D7、4C12、6G3
IgG2aκ	9D11、8F7
IgGMκ	2A5

杂交瘤细胞Hybridoma cell	3D7	4C12	6G3	9D11	8F7	2A5
3D7	-	28.68%	25.31%	20.26%	18.45%	25.92%
4C12	28.68%	-	37.69%	40.7%	29.98%	44.82%
6G3	25.31%	37.69%	-	23.6%	34.42%	15.53%
9D11	20.26%	40.7%	23.6%	-	13.63%	7.92%
8F7	18.45%	29.98%	34.42%	13.63%	-	28.54%
2A5	25.92%	44.82%	15.53%	7.92%	28.54%	-

项目 Item	双抗体夹心ELISA Double antibody sandwich ELISA	IBDV抗原检测卡 IBDV antigen test card
阳性样品数量 Number of positive samples	16	17
阴性样品数量 Number of negative samples	33	32

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IBDV单克隆抗体的制备及双抗体夹心ELISA检测方法的建立

Preparation of Monoclonal Antibodies to IBDV and Establishment of a Double Antibody Sandwich ELISA Method for Detection

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