纳米金比色法在家蚕微孢子虫检测中的应用
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摘要
家蚕微粒子病被列为蚕种生产的唯一检疫对象,母蛾镜检家蚕微孢子虫(Nosema bombycis,简称Nb)是目前最主要的检疫手段,但其准确性受到多种因素影响。建立一种简便、快捷、准确的Nb检测技术,对家蚕微粒子病的防治具有重要意义。基于纳米金(AuNPs)比色法原理,对AuNPs探针的制备及其标记的最佳抗体浓度和最佳pH值进行探究,以及探究AuNPs比色法检测Nb的灵敏度。结果表明,Nb抗体与AuNPs(20 nm)结合最佳浓度为每500μL AuNPs溶液中添加5μL 0.5 mg/mL抗体蛋白,标记最佳pH值为8.5。AuNPs比色法能够最低检测出Nb蛋白量为10 ng。本研究成功建立一种基于AuNPs比色法检测Nb的新方法。
引文
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[2]戴卫江,陈功,彭祥然,等.应用荧光染色剂Fluorescent Brightener 28和Propidium Iodide染色识别家蚕微孢子虫[J].蚕业科学,2017,43(1):62-67.
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[4]芦琨,党晓群,潘国庆,等.家蚕微孢子虫诊断方法研究进展[J].蚕学通讯,2008,28(4):35-38.
[5]刘吉平,曹阳,Smith J E,等.模拟感染家蚕微粒子病的PCR分子诊断技术研究[J].中国农业科学,2004,37(12):1925-1931.
[6]万淼,何永强,张海燕,等.家蚕成品卵微粒子病McAb-ELISA检测方法的研究[J].蚕桑通报,2008,39(4):13-16.
[7]芦琨.家蚕微粒子病免疫学快速检测试纸条的研制[D].重庆:西南大学,2009:35-43.
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[11]韦亚东,张国政,陆长德.家蚕微孢子虫原位杂交诊断技术研究[J].蚕业科学,2005,31(1):64-68.
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[13]Cordray M S,Amdahl M,Richards-Kortum R R .Gold nanoparticle aggregation for quantification of oligonucleotides:Optimization and increased dynamic range[J].Analytical Biochemistry,2012,431(2):99-105.
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[16]Jaroenram W,Arunrut N,Kiatpathomchai W.Rapid and sensitive detection of shrimp yellow head virus using loop-mediated isothermal amplification and a colorogenicnanogold hybridization probe[J].Journal of Virological Methods,2012,186(1/2):36-42.
[17]Shyu R H,Shyu H F,Liu H W,et al.Colloidal gold-based immunochromatographic assay for detection of ricin[J].Toxicon,2002,40(3):255-258.
[18]Zambre A,Chanda N,Prayaga S,et al.Design and development of a field applicable gold nanosensor for the detection of luteinizing hormone[J].Analytical Chemistry,2012,84(21):9478-9484.
[19]Guo A L,Sheng H L,Zhang M,et al.Development and evaluation of a colloidal gold immunochromatography strip for rapid detection of vibrio parahaemolyticus in food[J].Journal of Food Quality,2012,35(5):366-371.
[20]Lee J S,Han M,Mirkin C.Colorimetric detection of mercuric ion(Hg2+)using DNA-functionalized gold nanoparticles[J].AngewandteChem,2007,46(22):4093-4096.
[21]Li Y S,Zhou Y,Meng X Y,et al.Gold nanoparticle aggregation-based colorimetric assay for β-casein detection in bovine milk samples[J].Food Chemistry,2014,162:22-26.
[22]Mirkin C A,Letsinger R L,Mucic R C,et al.A DNA-based method for rationally assembling nanoparticles into macroscopic materials[J].Nature,1996,382(6592):607-609.
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[24]Burns C,Spendel W U,Puckett S,et al.Solution Ionic strength effect on gold nanoparticle solution color transition[J].Talanta,2006,69(4):873-876.
[25]He G,Zhao L,Chen K,et al.Highly selective and sensitive gold nanoparticle-based colorimetric assay for PO43- in aqueous solution[J].Talanta,2013,106(6):73-78.
[26]Zhou Y,Pan F G,Li Y S,et al.Colloidal gold probe-based immunochromatographic assay for the rapid detection of brevetoxins in fishery product samples[J].Biosensors & Bioelectronics,2009,24(8):2744-2747.
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[28]Li J,Wu Z,Wang H,et al.A reusable capacitive immunosensor with a novel immobilization procedure based on 1,6-hexanedithiol and nano-Au self-assembled layers[J].Sensors & Actuators (B:Chemical),2005,110(2):327-334.
[29]王伯阳,姜义仁,臧敏,等.柞蚕微孢子虫胶体金免疫层析检测法[J].蚕业科学,2012,38(1):97-101.