O型口蹄疫Cathay拓扑型病毒单抗制备及双抗体夹心ELISA方法的初步建立

doi:10.11843/j.issn.0366-6964.2024.09.025

Abstract

Abstract:

This study aimed to purify the cathay topotype of foot-and-mouth disease virus (FMDV) type O and prepare monoclonal antibody to provide biological materials for the study of novel cathay strain of foot-and-mouth disease. The antigen of cathay strain of FMDV type O was purified by PEG precipitation method and immunized BALB/c mice. Hybridoma cells were prepared by fusion of spleen cells and SP2/0 cells. Positive cells were screened by indirect ELISA and subcloned by limited dilution method. Two hybridoma cells, named 10E6 and 11C7, which were able to secrete monoclonal antibodies specifically against O-type FMDV Cathay strain were obtained. Additivity assay showed that the additivity rate of the two mAbs was 49.45%, and different epitopes were identified.Indirect ELISA and IFA showed that the two strains had good reactivity with O-type FMDV Cathay strain. The results of monoclonal antibody detection showed that the two monoclonal antibodies could specifically recognize Cathay FMDV strains and did not cross-react with other FMDV strains. The results of antibody subtype identification showed that the light chain of 10E6 monoclonal antibody was Kappa chain, the light chain of 11C7 monoclonal antibody was Lamda chain, and the heavy chain type of both strains was IgG2a. The results showed that two monoclonal antibodies specifically binding to O-type FMDV Cathay topological strain were successfully prepared in this study. Both monoclonal antibodies had good reactivity and specificity. Two monoclonal antibodies were used to establish a double antibody sandwich ELISA method for detecting O-type FMDV Cathay strain. The ELISA method had good specificity, repeatability and sensitivity, which laid a foundation for the rapid diagnosis, control and research of O-type foot and mouth disease virus Cathay strain.

Key words: foot-and-mouth disease virus type O, Cathay topotype, hybridoma cells, monoclonal antibody, ELISA

CLC Number:

Huancheng LIAO, Zhengwang SHI, Juncong LUO, Wanying WANG, Lu FENG, Jing ZHOU, Fan ZHANG, Xintai SHI, Hong TIAN. Preparation of Monoclonal Antibody against Cathay Topotype of FMDV Type O and Development of Double Antibody Sandwich ELISA for Cathay Topotype of FMDV Type O[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(9): 4012-4020.

Figures/Tables 7

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References 24

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OD_{450 nm}			AI/%
A_10E6	A_11C7	A_10E6+11C7	AI/%
1.73	1.91	2.72	49.45

项目 Item	O/Cathay	O/MYA98	AF/72	A/WH/09	对照组 Control
OD_{450 nm}	2.241	0.169	0.121	0.120	0.124
S/N	18.073	1.363	0.976	0.968	/

病毒悬液 Viral suspension	稀释倍数 Dilution ratio	批内重复性 Intra batch repeatability		批间重复性 Inter batch repeatability
病毒悬液 Viral suspension	稀释倍数 Dilution ratio	$\bar x \pm s$	变异系数/% CV	$\bar x \pm s$	变异系数/% CV
阳性样品	8×	16.19±0.27	1.68	16.02±0.35	2.18
Positive samples	16×	12.26±0.06	0.52	12.56±0.16	1.27
	32×	8.52 ± 0.42	4.94	8.12±0.52	6.40
阴性样品	8×	1.12±0.06	5.90	1.02±0.07	7.45
Negative samples	16×	0.95±0.06	6.71	0.93±0.07	7.93
	32×	0.68±0.05	7.53	0.70±0.06	8.73

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Preparation of Monoclonal Antibody against Cathay Topotype of FMDV Type O and Development of Double Antibody Sandwich ELISA for Cathay Topotype of FMDV Type O

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