Development of a Colloidal Gold Immunochromatographic Test Strip for Detection of Potato Virus X， Virus Y and Virus S

doi:10.13304/j.nykjdb.2021.0036

Abstract

Abstract:

To develop a colloidal gold immunochromatographic test strip for on-site rapid detection of potato （Solanum tuberosum L.） virus X （PVX）， virus Y （PVY） and virus S （PVS）， three-fold （PVX-PVY-PVS） test strip was prepared by colloidal gold labeling and immunochromatography. Colloidal gold solution was prepared by trisodium citrate reduction method. Gold-labeled antibody， which was composed of gold atom and PVX， PVY and PVS antibody， was prepared by the combination of antibody and colloidal gold. The gold-labeled antibody was sprayed on the glass fiber by a three-dimensional film-spraying gold-spraying instrument， which was used as a gold-labeled pad. In the strip test，colloidal gold-labeled antibodies against PVX， PVY and PVS were sprayed on the glass fiber as the detection antibody， and rabbit anti-goat IgG at the control line， anti-PVX， anti-PVY and anti-PVS at the test line on the nitrocellulose membrane of the test strip were served as the capture antibody. The results showed that the three kinds of viruses （PVX， PVY and PVS） could be simultaneously detected within 5 min by the triple test strip .The detection was still feasible when the PVX samples was diluted by 10⁴ （W/V），the PVY samples was diluted by 2×10³ （W/V）， the PVS samples was diluted by 10³ （W/V）， which was more sensitive to PVX detection. The triple test strip showed no cross effects to common 3 viruses （PLRV， PVA and PVM） samples. The results of triple test strip on potato leaves collected in the field were consist with DAS-ELISA. The triple test strip had the characteristics of simple operation and quick reaction， which should be suitable for field and port inspection especially.

Key words: potato virus X, potato virus Y, potato virus S, triple test strip

摘要：

为实现在马铃薯种薯生产田现场同时快速检测马铃薯X病毒（PVX）、Y病毒（PVY）和S病毒（PVS），采用柠檬酸三钠还原法制备胶体金溶液，利用抗体与胶体金相结合的特性制备金原子与PVX、PVY和PVS抗体结合物（简称金标抗体），通过三维划膜喷金仪将金标抗体喷射于玻璃纤维上作为金标垫，将PVX抗体、PVY抗体和PVS抗体分别划线于硝酸纤维素膜作为检测线，将羊抗兔抗体划线于硝酸纤维素膜作为质控线，进行三重胶体金试纸条的制备。结果表明，研制的三重试纸条能够在5 min内同时检出PVX、PVY和PVS 3种病毒。PVX病汁液检测稀释限度可达10⁴倍（W/V），PVY病汁液检测稀释限度可达2×10³倍（W/V），PVS病汁液检测稀释限度可达10³倍（W/V），即对PVX检测更灵敏。利用研制的三重检测试纸条检测危害马铃薯的其他3种常见病毒，未出现交叉反应。对田间采集的马铃薯叶片进行检测，发现利用试纸条与酶联免疫吸附法（DAS-ELISA）检测的结果完全一致。三重检测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的检测。

关键词: 马铃薯X病毒, 马铃薯Y病毒, 马铃薯S病毒, 三重检测试纸条

CLC Number:

S532

Wei ZHANG, Zhixin LI, Xue ZHAO, Jinpeng ZHANG, Chunjiang FU, Weiping LIU, Qianqian YU. Development of a Colloidal Gold Immunochromatographic Test Strip for Detection of Potato Virus X， Virus Y and Virus S[J]. Journal of Agricultural Science and Technology, 2022, 24(1): 211-217.

张微, 李志新, 赵雪, 张金鹏, 付春江, 刘卫平, 于倩倩. 同时快速检测马铃薯X病毒、Y病毒和S病毒试纸条的研制[J]. 中国农业科技导报, 2022, 24(1): 211-217.

Figures/Tables 7

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pH	K₂CO₃/µL	1	2	3	4	5	6	7	8
pH	颜色 Color	酒红色Claret	酒红色Claret	红色Red	红色Red	红色Red	红色Red	红色Red	红色Red
抗体 Antibody	用量Dosage/µL	1	2	3	4	5	6	7	8
抗体 Antibody	沉淀Precipitate	无Nothing	无Nothing	无Nothing	无Nothing	极少量 Very few	极少量 Very few	少量 Few	少量Few
抗体 Antibody	加入顺序Order	PVX、PVY、PVS	PVX、PVS、PVY	PVY、PVX、PVS	PVY、PVS、PVX	PVS、PVX、PVY	PVS、PVY、PVX
抗体 Antibody	颜色Color	酒红色Claret	酒红色Claret	红色Red	粉红色Pink	淡紫色Lavender	淡紫色Lavender
封闭 Closing	时间Time/min	20	25	30	35	40	45	50
封闭 Closing	颜色Color	红色Red	红色Red	酒红色Claret	酒红色Claret	酒红色Claret	酒红色Claret	酒红色Claret
离心 Centrifugal	时间Time/min	20	25	30	35	40	45	50
离心 Centrifugal	沉淀 Precipitate	吸上清，易悬浮 Absorb supernatant， easy to suspend	吸上清，易悬浮 Absorb supernatant， easy to suspend	吸上清，不易悬浮 Absorb supernatant， not easy to suspend	吸上清，不易悬浮 Absorb supernatant， not easy to suspend	吸上清，不易悬浮 Absorb supernatant， not easy to suspend	吸上清，不易悬浮Absorb supernatant， not easy to suspend	吸上清，不易悬浮Absorb supernatant， not easy to suspend
复溶液 Complex solution	用量Dosage/µL	80	90	100	110	120
复溶液 Complex solution	显色 Color	深红色Oxblood red	深红色Oxblood red	酒红色Claret	红色 Red	粉红色 Pink

pH	K₂CO₃/µL	1	2	3	4	5	6	7	8
pH	颜色 Color	酒红色Claret	酒红色Claret	红色Red	红色Red	红色Red	红色Red	红色Red	红色Red
抗体 Antibody	用量Dosage/µL	1	2	3	4	5	6	7	8
抗体 Antibody	沉淀Precipitate	无Nothing	无Nothing	无Nothing	无Nothing	极少量 Very few	极少量 Very few	少量 Few	少量Few
抗体 Antibody	加入顺序Order	PVX、PVY、PVS	PVX、PVS、PVY	PVY、PVX、PVS	PVY、PVS、PVX	PVS、PVX、PVY	PVS、PVY、PVX
抗体 Antibody	颜色Color	酒红色Claret	酒红色Claret	红色Red	粉红色Pink	淡紫色Lavender	淡紫色Lavender
封闭 Closing	时间Time/min	20	25	30	35	40	45	50
封闭 Closing	颜色Color	红色Red	红色Red	酒红色Claret	酒红色Claret	酒红色Claret	酒红色Claret	酒红色Claret
离心 Centrifugal	时间Time/min	20	25	30	35	40	45	50
离心 Centrifugal	沉淀 Precipitate	吸上清，易悬浮 Absorb supernatant， easy to suspend	吸上清，易悬浮 Absorb supernatant， easy to suspend	吸上清，不易悬浮 Absorb supernatant， not easy to suspend	吸上清，不易悬浮 Absorb supernatant， not easy to suspend	吸上清，不易悬浮 Absorb supernatant， not easy to suspend	吸上清，不易悬浮Absorb supernatant， not easy to suspend	吸上清，不易悬浮Absorb supernatant， not easy to suspend
复溶液 Complex solution	用量Dosage/µL	80	90	100	110	120
复溶液 Complex solution	显色 Color	深红色Oxblood red	深红色Oxblood red	酒红色Claret	红色 Red	粉红色 Pink