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基于磁珠纯化的聚合酶链反应结合荧光侧向流动免疫分析法用于快速检测犬细小病毒2型

Polymerase chain reaction combined with fluorescent lateral flow immunoassay based on magnetic purification for rapid detection of canine parvovirus 2.

作者信息

Zhuang Linlin, Ji Yongxin, Tian Peilong, Wang Kaixuan, Kou Chengkun, Gu Ning, Zhang Yu

机构信息

State Key Laboratory of Bioelectronics, Jiangsu Key Laboratory for Biomaterials and Devices, School of Biological Sciences and Medical Engineering and Collaborative Innovation Center of Suzhou Nano Science and Technology, Southeast University, No. 2, Sipailou, Xuanwu District, Nanjing, Jiangsu Province, 210096, People's Republic of China.

Nanjing Nanoeast Biotech Co., Ltd., Nanjing, Jiangsu, 210009, People's Republic of China.

出版信息

BMC Vet Res. 2019 Jan 17;15(1):30. doi: 10.1186/s12917-019-1774-3.

DOI:10.1186/s12917-019-1774-3
PMID:30654823
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC6337814/
Abstract

BACKGROUND

Canine parvovirus 2 (CPV-2) is one of the most common etiological agents that cause severe gastroenteritis in puppies. Early accurate diagnosis is important for infected dogs. In recent years, magnetic separation has become an efficient and useful tool for bioassays. In this study, polymerase chain reaction (PCR) combined with fluorescent lateral flow immunoassay (LFIA) based on magnetic purification assay was developed for the quantitative detection of CPV-2.

RESULTS

The optimum working reaction volume and reaction time for LFIA was 100 μL and 2 min, respectively. The PCR-LFIA assay only detected CPV-2, and did not show cross-detection of non-CPV strains. Experiments showed analytical sensitivity of 3 × 10 copies/μL and demonstrated the PCR-LFIA has a diagnostic agreement of 100% with conventional PCR on detection of clinical samples (22.6% positive, 14/62). Cutoff value is 146. The results were further verified by sequencing and BLAST software. The entire process from PCR step only takes ~ 80 min.

CONCLUSIONS

This approach provides an attractive platform for rapid and quantitative detection of CPV-2, indicating great promise as a convenient molecular detection tool to facilitate disease outbreak investigations and response timely.

摘要

背景

犬细小病毒2型（CPV - 2）是引起幼犬严重肠胃炎的最常见病原体之一。早期准确诊断对感染犬至关重要。近年来，磁分离已成为生物检测的一种高效且有用的工具。在本研究中，基于磁纯化分析开发了聚合酶链反应（PCR）结合荧光侧向流动免疫分析（LFIA）用于CPV - 2的定量检测。

结果

LFIA的最佳工作反应体积和反应时间分别为100 μL和2分钟。PCR - LFIA分析仅检测CPV - 2，未显示对非CPV毒株的交叉检测。实验显示分析灵敏度为3×10拷贝/μL，并且在检测临床样本（22.6%阳性，14/62）时，PCR - LFIA与传统PCR的诊断一致性为100%。临界值为146。结果通过测序和BLAST软件进一步验证。仅PCR步骤的整个过程仅需约80分钟。

结论

该方法为CPV - 2的快速定量检测提供了一个有吸引力的平台，显示出作为一种方便的分子检测工具促进疾病爆发调查和及时应对的巨大潜力。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/22c4e2702c82/12917_2019_1774_Fig7_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/ebd05e461417/12917_2019_1774_Fig1_HTML.jpg
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https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/e76879fc2e4b/12917_2019_1774_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/8faf6843d34b/12917_2019_1774_Fig6_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/22c4e2702c82/12917_2019_1774_Fig7_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/ebd05e461417/12917_2019_1774_Fig1_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/7dacb34df85f/12917_2019_1774_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/63223ddba333/12917_2019_1774_Fig3_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/f4922e87bd99/12917_2019_1774_Fig4_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/e76879fc2e4b/12917_2019_1774_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/8faf6843d34b/12917_2019_1774_Fig6_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1bec/6337814/22c4e2702c82/12917_2019_1774_Fig7_HTML.jpg

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