Development and preliminary application of a one-step RT-qPCR method for de-tection of porcine epidemic diarrhea virus based on immunomagnetic beads
10.16303/j.cnki.1005-4545.2025.09.01
- VernacularTitle:基于免疫磁珠检测猪流行性腹泻病毒的一步RT-qPCR方法的建立及初步应用
- Author:
Shunli YANG
1
;
Xiaojuan LYU
;
Li LI
;
Xiaoqing ZHANG
;
Yupeng FANG
;
Tian ZHAO
;
Jiqiao XIA
;
Jie ZHANG
;
Zhixin FU
;
Yongsheng LIU
Author Information
1. 河北科技师范学院动物科技学院河北省预防兽医学重点实验室,河北秦皇岛 066500
- Publication Type:Journal Article
- Keywords:
porcine epidemic diarrhea virus;
nanobody;
immunomagnetic beads;
reverse transcriptase polymerase chain reaction
- From:
Chinese Journal of Veterinary Science
2025;45(9):1817-1823
- CountryChina
- Language:Chinese
-
Abstract:
In this study,the carboxylated magnetic beads were coupled with bivalent nanobodies a-gainst porcine epidemic diarrhea virus(PEDV)M protein to construct immunomagnetic beads(IM-NBs-Ⅱ),The capture and enrichment function of IMNBs-Ⅱ was verified by using PEDV propaga-ted in Vero cells.A one-step RT-qPCR detection method for PEDV was established by combining the characteristics of IMNBs-Ⅱ with the detection advantages of reverse transcription fluorescence quantitative polymerase chain reaction(RT-qPCR).Specific analysis found that this method has no cross reactivity with swine fever virus,porcine reproductive and respiratory syndrome virus,por-cine parvovirus,porcine circovirus,indicating that it has good specificity.Sensitivity analysis re-vealed that the detection sensitivity of the RT-qPCR based on IMNBs-Ⅱ was increased 10 times compared to traditional RT-qPCR methods.Detection of the clinical samples confirm that the RT qPCR method based on IMNBs-Ⅱ is suitable for rapid and accurate detection of clinical feces and tissue samples.The method established in this study effectively avoids contamination issues during nucleic acid extraction,simplifies experimental procedures,and saves detection time,which pro-vides a method for efficient detection of PEDV.
