贝类腹泻性毒素现场高通量前处理及快速检测仪器设计
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摘要
为实现贝类腹泻性毒素的现场快速检测,本论文设计了一种现场高通量前处理装置及基于移动终端的快速检测系统。通过使用酶联免疫反应技术对腹泻性贝毒进行显色反应,结合基于移动终端的快速检测装置,采集显色反应结果图像,实现准确快速的检测结果分析。采用所设计的前处理装置与实验室手工前处理标准方法对比,其回收率分别为89%和93%,结果表明,本文设计的前处理装置能够满足现场检测分析的需求。通过加标样品检测,采用移动终端的快速检测系统测量标准差为0.13,平均回收率89.5%。实验结果表明,该检测系统的准确性和重复性满足实际检测的需求。最后,针对实际样品,与酶标仪检测结果相比较,表明了所设计的现场高通量前处理装置及快速检测系统能达到对贝类腹泻性毒素现场快速精确检测,为贝类腹泻性毒素现场检测提供了新的方法和仪器。
In order to achieve the requirement of diarrhetic shellfish poison( DSP) in-suit detection,we design a high throughput pretreatment system and a mobile terminal based detection system. Our system can make fast and accurate DSP detection by analyzing image of chromogenic reaction come from ELISA method. The rate of recovery by our pretreatment system and by hand are 89% and 93% respectively,it means that our pretreatment system can meet the follow-up detection requirement. Detect the prevented samples by our detection system,the standard deviation is approximately 0.13 and average rate of recovery is 89.5%,it can satisfy the accuracy and repeatability of detection. In the detection of actual samples,comparing our results with the results come from microplate reader,it shows that our pretreatment and detection system can do a fast and accurate in-suit work and provide a new method for the detection.
In order to achieve the requirement of diarrhetic shellfish poison( DSP) in-suit detection,we design a high throughput pretreatment system and a mobile terminal based detection system. Our system can make fast and accurate DSP detection by analyzing image of chromogenic reaction come from ELISA method. The rate of recovery by our pretreatment system and by hand are 89% and 93% respectively,it means that our pretreatment system can meet the follow-up detection requirement. Detect the prevented samples by our detection system,the standard deviation is approximately 0.13 and average rate of recovery is 89.5%,it can satisfy the accuracy and repeatability of detection. In the detection of actual samples,comparing our results with the results come from microplate reader,it shows that our pretreatment and detection system can do a fast and accurate in-suit work and provide a new method for the detection.
引文
[1]张少君,丁永生,李大志,等.贝毒及检测方法的研究进展[J].大连海事大学学报:自然科学版,2004,29(4):62-65.
[2]Haystead T A J,Sim A,Carling D,et al.Effects of the Tumour Promoter Okadaic Acid on Intracellular Protein Phosphorylation and Metabolism[J].Nature,1989,337(6202):78-81.
[3]Stookey D,Patton C,Malcolm G.Membrane Separation Technology.American Institute of Chemical Engineers,New York,NY,1986.
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[7]曹际娟,卫锋,马惠蕊,等.贝类毒素检测技术及研究进展[J].检验检疫科学,2004,14(1):53-56.
[8]Carmichael W,Evans W,Yin Q,et al.Evidence for Paralytic Shellfish Poisons in the Freshwater Cyanobacterium Lyngbya Wollei(Farlow ex Gomont)Comb.Nov[J].Applied and Environmental Microbiology,1997,63(8):3104-3110.
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[10]黄玉柳,黄国秋,叶欣宇,等.酶联免疫法快速检测贝类中腹泻性贝类毒素[J].化学与生物工程,2011,28(11):93-94.
[11]王峰,刘飞,孟令花,等.基于抗体包被金磁纳米微粒修饰的磁性安培免疫传感器研制及对人血清癌抗原19-9的检测[J].传感技术学报,2009(9):1232-1238.
[12]Usleber E,Dietrich R,Bürk C,et al.Immunoassay Methods for Paralytic Shellfish Poisoning Toxins[J].Journal of AOAC International,2001,84(5):1649-1656.
[13]Qi-Fang L.Detection of Paralytic Shellfish Poisoning(PSP),Diarrhetic Shellfish Poison(DSP),Neurotoxic Shellfish Poison(NSP)by ELISA[J].China Tropical Medicine,2012,11:38.
[14]苏凯麒,邹玲,王琴,等.基于细胞阻抗传感器的腹泻性毒素检测系统设计与实现[J].传感技术学报,2014,27(3):283-288.
[15]谢东华,干宁,王峰,等.鱼肉中氯霉素检测用抗体包被金磁纳米微粒修饰安培免疫传感器[J].传感技术学报,2009(10):1371-1377.
[16]Vashist S K,van Oordt T,Schneider E M,et al.A SmartphoneBased Colorimetric Reader for Bioanalytical Applications Using the Screen-Based Bottom Illumination Provided by Gadgets[J].Biosensors and Bioelectronics,2015,67:248-255.
[17]Berg B,Cortazar B,Tseng D,et al.Cellphone-Based Hand-Held Microplate Reader for Point-of-Care Testing of Enzyme-Linked Immunosorbent Assays[J].ACS Nano,2015,9(8):7857-7866.
[18]Dixon L G,Hamdy M K,Hayden M A,et al.Elisa for Detecting Okadaic Aced in Model Systems Using Purified Polyclonal Antibodies[J].Journal of Rapid Methods and Automation in Microbiology,1997,5(4):269-283.
[19]Cusick K D,Sayler G S.An Overview on the Marine Neurotoxin,Saxitoxin:Genetics,Molecular Targets,Methods of Detection and Ecological Functions[J].Marine Drugs,2013,11(4):991-1018.
[20]Rossini G P.Functional Assays in Marine Biotoxin Detection[J].Toxicology,2005,207(3):451-462.
[2]Haystead T A J,Sim A,Carling D,et al.Effects of the Tumour Promoter Okadaic Acid on Intracellular Protein Phosphorylation and Metabolism[J].Nature,1989,337(6202):78-81.
[3]Stookey D,Patton C,Malcolm G.Membrane Separation Technology.American Institute of Chemical Engineers,New York,NY,1986.
[4]Hennion M C.Solid-Phase Extraction:Method Development,Sorbents,and Coupling with Liquid Chromatography[J].Journal of Chromatography A,1999,856(1):3-54.
[5]Hummert C,Shen J,Luckas B.Automatic High-Performance Liquid Chromatographic Method for the Determination of Diarrhetic Shellfish Poison[J].Journal of Chromatography A,1996,729(1):387-392.
[6]Ben-Gigirey B,Rodríguez-Velasco M,Otero A,et al.A Comparative Study for PSP Toxins Quantification by Using MBA and HPLC Official Methods in Shellfish[J].Toxicon,2012,60(5):864-873.
[7]曹际娟,卫锋,马惠蕊,等.贝类毒素检测技术及研究进展[J].检验检疫科学,2004,14(1):53-56.
[8]Carmichael W,Evans W,Yin Q,et al.Evidence for Paralytic Shellfish Poisons in the Freshwater Cyanobacterium Lyngbya Wollei(Farlow ex Gomont)Comb.Nov[J].Applied and Environmental Microbiology,1997,63(8):3104-3110.
[9]吴施卫,张纯超,卢楚谦,等.南海近岸海域腹泻性贝类毒素分析[J].海洋环境科学,2005,24(4):48-51.
[10]黄玉柳,黄国秋,叶欣宇,等.酶联免疫法快速检测贝类中腹泻性贝类毒素[J].化学与生物工程,2011,28(11):93-94.
[11]王峰,刘飞,孟令花,等.基于抗体包被金磁纳米微粒修饰的磁性安培免疫传感器研制及对人血清癌抗原19-9的检测[J].传感技术学报,2009(9):1232-1238.
[12]Usleber E,Dietrich R,Bürk C,et al.Immunoassay Methods for Paralytic Shellfish Poisoning Toxins[J].Journal of AOAC International,2001,84(5):1649-1656.
[13]Qi-Fang L.Detection of Paralytic Shellfish Poisoning(PSP),Diarrhetic Shellfish Poison(DSP),Neurotoxic Shellfish Poison(NSP)by ELISA[J].China Tropical Medicine,2012,11:38.
[14]苏凯麒,邹玲,王琴,等.基于细胞阻抗传感器的腹泻性毒素检测系统设计与实现[J].传感技术学报,2014,27(3):283-288.
[15]谢东华,干宁,王峰,等.鱼肉中氯霉素检测用抗体包被金磁纳米微粒修饰安培免疫传感器[J].传感技术学报,2009(10):1371-1377.
[16]Vashist S K,van Oordt T,Schneider E M,et al.A SmartphoneBased Colorimetric Reader for Bioanalytical Applications Using the Screen-Based Bottom Illumination Provided by Gadgets[J].Biosensors and Bioelectronics,2015,67:248-255.
[17]Berg B,Cortazar B,Tseng D,et al.Cellphone-Based Hand-Held Microplate Reader for Point-of-Care Testing of Enzyme-Linked Immunosorbent Assays[J].ACS Nano,2015,9(8):7857-7866.
[18]Dixon L G,Hamdy M K,Hayden M A,et al.Elisa for Detecting Okadaic Aced in Model Systems Using Purified Polyclonal Antibodies[J].Journal of Rapid Methods and Automation in Microbiology,1997,5(4):269-283.
[19]Cusick K D,Sayler G S.An Overview on the Marine Neurotoxin,Saxitoxin:Genetics,Molecular Targets,Methods of Detection and Ecological Functions[J].Marine Drugs,2013,11(4):991-1018.
[20]Rossini G P.Functional Assays in Marine Biotoxin Detection[J].Toxicology,2005,207(3):451-462.