金纳米粒子增强免疫 PCR 法高灵敏检测口蹄疫病毒
A highly sensitive detection for foot-and-mouth disease virus by gold nanopariticle improved immuno-PCR.
机构信息
Chinese Academy of Agricultural Sciences, Lanzhou, Gansu,China.
出版信息
Virol J. 2011;8:148. doi: 10.1186/1743-422X-8-148. Epub 2011 Mar 31.
BACKGROUND
Foot-and-mouth disease (FMD) is one of the most contagious of all artiodactyl animal diseases, and its infection has an obvious ability to spread over long distances and to contribute to epidemics in FMD-free areas. A highly sensitive and specific method is required to detect FMDV. In this study, we evaluated the usefulness of a bio-barcode assay (BCA) technique for detecting clinical samples of FMDV.
METHODS
Highly sensitive gold nanopariticle (GNP) improved immuno -PCR (GNP-IPCR) which derived from the bio-barcode assay (BCA) was designed for the detection of FMDV. The target viral particles were captured by a polyclonal antibody coated on ELISA microplate, followed by adding GNP which was dually modified with oligonucleotides and a FMDV specific monoclonal antibody (MAb) 1D11 to form a sandwiched immune complex. After the formation of immuno-complex, the signal DNA was released by heating, and consequently characterized by PCR and real time PCR.
RESULTS
The detection limit of GNP-PCR could reach to 10 fg/ml purified FMDV particles, and the assay can detect clinical samples of FMDV with highly sensitivity, while detect limit of conventional ELISA is 100 ng/ml in this study.
CONCLUSION
GNP-IPCR may provide a highly sensitive method for the detection of FMDV.
背景
口蹄疫（FMD）是所有偶蹄动物疾病中最具传染性的疾病之一，其感染具有明显的远距离传播能力，并有助于无口蹄疫地区的流行。需要一种高度敏感和特异的方法来检测 FMDV。在本研究中，我们评估了生物条码检测技术（BCA）在检测 FMDV 临床样本中的应用价值。
方法
高灵敏度的金纳米粒子（GNP）改良免疫-PCR（GNP-IPCR）来源于生物条码检测技术（BCA），用于检测 FMDV。目标病毒颗粒通过包被在 ELISA 微孔板上的多克隆抗体捕获，然后加入双重修饰有寡核苷酸和 FMDV 特异性单克隆抗体（MAb）1D11 的 GNP，形成夹心免疫复合物。免疫复合物形成后，通过加热释放信号 DNA，随后通过 PCR 和实时 PCR 进行特征分析。
结果
GNP-PCR 的检测限可达到 10 fg/ml 纯化的 FMDV 颗粒，该方法具有高度的灵敏度，可检测临床 FMDV 样本，而本研究中常规 ELISA 的检测限为 100ng/ml。
结论
GNP-IPCR 可能为 FMDV 的检测提供一种高度敏感的方法。
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