双重检测马铃薯X和Y病毒试纸条制备技术研究

为了实现在马铃薯种薯生产田现场同时快速检测马铃薯X（PVX）和Y病毒（PVY）,利用胶体金标记和免疫层析技术研制了PVX-PVY双重检测试纸条。采用柠檬酸三钠还原法制备胶体金溶液,在波长526nm处有最大吸收值,金颗粒直径约25nm。同时标记PVX和PVY兔多克隆抗体,将抗体标记物喷射于玻璃纤维纸上,作为金标垫。将PVX和PVY抗体分别划线于硝酸纤维素膜上作为检测线,将羊抗兔抗体划线于硝酸纤维素膜上作为质控线,制备胶体金试纸条。结果表明,研制的双重试纸条能够在2min之内同时检出PVX和PVY。PVX病汁液检测稀释限度可达10⁶倍（W/V）,PVY可达10⁵倍（W/V）,其中对PVX检测更灵敏。用其他4种常见病毒（PVS、PLRV、PVA和PVM）检测,未出现交叉反应。在田间采集马铃薯叶片,分别利用试纸条与DAS-ELISA检测,结果一致性非常高。由此可知,该试纸条具有操作简便、反应快捷的特点,特别适用于田间及口岸一线对于马铃薯X和Y病毒的同时检测。

Development of a Double Test Strip for the Detection of Potato Virus X and Y

In order to detect potato virus X (PVX) and Y (PVY) rapidly in potato seed production field, a PVX-PVY test strip was developed by colloidal gold labeling and immunochromatography. Colloidal gold solution was prepared by trisodium citrate reduction method with the maximum absorption value at the wavelength of 526nm and the diameter of gold particles was about 25nm. In the strip test, colloidal gold-labeled polyclonal rabbit antibody against PVX and PVY were sprayed on the glass fiber as the detection antibody, and goat anti-rabbit IgG at the control line and anti-PVX/ anti-PVY at the test line on the nitrocellulose membrane of the test strip were served as the capture antibody. The results showed that the two kinds of viruses could be simultaneously detected within 2min. The detection was still feasible when the PVX sample was diluted by 10⁶(W/V), the PVY sample was diluted by 10⁵ (W/V), which was more sensitive to PVX detection. The test strip showed no cross effect when tested by other common four viruses (PVS, PLRV, PVA, and PVM) samples. DAS-ELISA was used to detect the consistency of potato leaves collected in the field. The strip has the characteristics of simple operation and quick reaction, especially suitable for field and port inspection.