摘要
本文以MCF-7细胞和PC-3细胞为模型细胞,采用电化学法、HPLC法、化学计量学法测定了细胞的伏安行为,分析了细胞电化学活性物质的种类和含量,研究和揭示了细胞电化学伏安响应机理;通过对比细胞分泌液和细胞质的伏安行为,建立了基于细胞质伏安行为的电化学检测环境雌激素活性新方法,使环境雌激素的筛选更有效、简单、快速和灵敏。研究结果概括如下:
1、揭示了细胞电化学响应机理。研究了活细胞悬浮液和细胞质的电化学行为,发现活细胞悬浮液的电化学信号来源于细胞分泌物中鸟嘌呤和黄嘌呤的氧化反应,而不是通常人们推测的来源于活细胞本体或酶的电化学响应。本文采用了超声波和加热灭活法破碎活细胞得到细胞质,并检测了其电化学信号,发现细胞质的电化学信号远高于活细胞悬浮液,并且加热灭活法获得的细胞质的伏安信号更强,这是由于该方法获得的细胞质中鸟嘌呤和黄嘌呤的含量较高。
2、建立了电化学法评价细胞活性系统。为了揭示细胞电化学信号与细胞活性的关系,本文采用了经典天然抗癌药物紫杉醇和系列非强效天然抗癌药物黄酮类化合物作为细胞活性抑制剂,使用电化学法研究了药物对细胞活性的影响。结果发现,随着药物作用时间和剂量的增加,细胞活性下降,细胞质中鸟嘌呤和黄嘌呤的浓度下降,细胞质电化学信号也随之下降,并具有时间和剂量依赖性。采用MTT和流式细胞仪常规检测法验证电化学法的可靠性,结果发现,两者的检测结果与电化学法基本一致,说明基于细胞质伏安行为建立的电化学评价细胞活性方法具有高灵敏性和可靠性。揭示了细胞质电化学行为与核苷酸代谢之间的关系。
3、建立了细胞电化学检测环境雌激素活性方法。比较了去激素MCF-7细胞与正常培养MCF-7细胞电化学行为及其细胞质中电活性物质成分的区别。结果发现,去激素MCF-7细胞质中的黄嘌呤和鸟嘌呤浓度降低,使细胞分泌黄嘌呤和鸟嘌呤能力降低,导致细胞和细胞质的伏安信号减弱,说明去激素MCF-7细胞的活性下降;研究了标准雌激素雌二醇、环境雌激素壬基酚和双酚A对去激素培养的MCF-7细胞增殖的影响,发现其均存在剂量-效应关系和时间-效应关系,并且雌激素效应由大到小的顺序为:雌二醇＞壬基酚＞双酚A;同时使用3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)法验证了电化学法评价雌激素效应的可靠性。结果发现,两者的检测结果基本一致,说明细胞电化学法检测环境雌激素活性是可行的。
This paper studies the voltammetric response of the MCF-7 cells and the PC-3 cells as the model cell by using electrochemical method, HPLC method and chemometric analysed the kinds and the concentration of the electrochemical active specises of the cells, and explained the mechanism on the origin of the voltammetric response; By comparison of the electrochemical behaviors of the cell secretion and the cytoplasm, the electrochemical method of measuring active of the environmental estrogens based on the electrochemical behaviors of the cytoplasm was built. The electrochemical method are rapid, highly sensitive, and inexpensive. The results are as follows:
1. The mechanism on the origin of the voltammetric response of the cells. The electrochemical behaviors of the living cell suspension and the cytoplasm were studied, and the results were found that the electrochemical response of the living cell suspension was given by the oxidation of guanine and xanthine in the cell secretion, not by the cells. Ultrasonication and heat treatment were carried out for obtaining the cytoplasm, whose voltammeric response was stronger than that of the cell suspension. The voltammeric response of the cytoplasm obtained by heat treatment was stronger than that by ultrasonication due to that the concentrations of guanine and xanthine in the cytoplasm obtained heat treatment were higher than that by ultrasonication.
2. Building the system of evaluating the active of the cells by the electrochemical method. To suggest the relation between the electrochemical signal and the cell viability, the effect of cytotoxicity of taxol, luteolin, quercetin, isoquercitin and rutin on the cell viability was studied by electrochemical method. The result obtained by using the electrochemical method displayed cytotoxic tendencies similar to those by the conventional in vitro drug MTT and flow cytometry sensitivity tests, suggesting that the method of recording voltammetric measurements described above was credible. The altered response was driven by the decreased concentration of xanthine and guanine in the cytoplasm. The relation between the voltammeric behavior of cells and intracellular catabolism was interprected.
3. Establishment the method of measuring environmental estrogens based on the cell electrochemistry. Comparative studies of electroactive components and voltammetric behaviors different between the MCF-7 cell without estrogen and the MCF-7 cell with estrogen showed that the concentration of the guanine and xanthine bases in the cytoplasm decreased, which resulted in the decrease of secretion ability of the MCF-7 cell without estrogen decreased, and these result in the decreasing of the voltammetric response of the cells, impling that the cell vaibility decreases. The effect of estradio. nonylphenol.bisphenol on the proliferation of the MCF-7 cells without estrogen.The results showed that dosage-dependen and time-dependen was present and the order of the estrogen effect is as follow: estradio>nonylphenol>bisphenol; The effect of environmental estrogens on the proliferation of the MCF-7 cells was studied by the electrochemica method, and the result above was similar to those by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method, suggesting that voltammetric measurements was credible.
引文
[1]Kavlock R J,Daston G P,Derosa C,et al.Research needs for the risk assessment of health and environment effects of endocrine disruptors:A report of the USEPA- sponsored workshop.Environ Health Perspect,1996,104:715-740.
[2]Ahmed S A.The immune system as a potential target for environmental estrogens endocrine disrupters:a new emerging field.Toxicology,2000,150:191-206.
[3]中华人民共和国卫生部.中国标准出版社.食品添加剂使用卫生标准 GB2760-1996[S].
[4]郯云雁.美国的食品添加剂管理[J].中国食品卫生杂志,1999,(2):77-79.
[5]邱东茹,吴振斌,贺锋.内分泌扰乱化学品对动物的影响和作用机制,环境科学研究,2000.1(6):25-55.
[6]Kavlock R J.Over view of endocrine disruptor research activity in the United States.Chemasphere,1999,39:1227-1236.
[7]Arnold F,Robinson M K,Notides A C,et al.A yeast estrogen screen for examining the relative exposure of cells to natural and xenoestrogens[J].Environ Health Perspect,1996,104(5):544-548.
[8]Lachlan M,Arnold J A,et al.Potency of Combined Estrogenic Pesticides,Science,1997,275(5298):405-406.
[9]Hayes T,Haston K.Tsui M,et al.Feminization of Male Frogs in the Wild.Nature,2002,419(6910):895-896.
[10]伊雄海.农药类环境激素低剂量暴露对鲫鱼分泌干扰效应及生物标忠物研究[D]:[博士论文].上海:上海交通大学,2008.
[11]McAvoy D C,Eckhoff W S,Rapaport R A,et al.Fate of linear aylkybenzene sulfonates in the environment[J].Toxic.Chem.,1993(12):977-987.
[12]袁俊峰,陈浩文,吴宝玲,等.南极麦克斯韦尔海湾阴离子洗涤剂污染研究[J].中国环境科学,1998.18(2):151-153.
[13]芮旻,徐斌,陈艺,等.饮用水中阴离子合成洗涤剂去除的研究[J].工业用水与废水,2005,36(5):20-23.
[14]杨颖,黄国兰,孙江文.烷基酚和烷基酚聚氧乙烯醚的环境行为[J].安全与环境学报,2005,5(6):38-42.
[15]Maki H,Okamura H,Aoyama I,et al.Halogenation and toxicity of the biodegradation products of a nonionic surfactant,nonyphenol ethoxylate[J].Environ Toxicol Chem,1998,179:630-634.
[16]White R,Jobling S,Hoare S A,et al.Environmental persistent alkylphenolic compounds are estrogenic[J].Endocrinology,1994(135):175-182.
[17]Ashfield L A,Pottinger T G,Sumpter J P.Exposure of female juvenile rainbow trout to alkylphenolic compounds results in modification to growth and ovosomatic index[J].Environ Toxicol Chem,1998(17):679-686.
[18]Isobe T,Nishiyama H,Nakashima A,et al.Distribution and behavior on onylphenol,octylphenol,and nonylphenoi monoethoxylate in Tokyo metropolitan area:their association with aquatic particles and sedimentary distributions[J].Environ Sci Technol,2001(35):1041-1049.
[19]Sole M,Lopez D A M,Castilio M.Estrogenicity determination in sewage treatment plants and surface waters from the Catalonian area(NE Spain)[J].Environ Sci Technol,2000(34):5076-5083.
[20]Ferguson P L,lden C R,Brownwell B.1.Distribution and fate of neutral alkylphenol ethoxylate metabolites in a sewage-impacted urban estuary[J].Environ Sci Technol,2001(35):2428-2435.
[21]Komori K,Okayasu Y,Yasojima M,et al.Occurrence of nonylphenol nonylphenoi ethoxylate surfactants and nonyl-phenol carboxylic acids in wastewater in Japan[J].Water Science and Technology,2006(53):27-33.
[22]Schmitt K,Hirose T,Kinj O J,et al.Interaetion of Phytoesogen sineells of different estrogen sensitive tissues.Toxieol In Vitro,2002(15):433-439.
[23]Jensen E V,Jaeobsen H L.Basic guides to the mechanism of estrogen action.Recent Prog Horm Res,1962(18):387-414.
[24]冯士清,赵华.从使用频率看化妆品防腐剂走势(续前)[J].日用化学品科学,2006,29(12):32-35.
[25]王鹏,李洁,朱杰.化妆品中七种对经基苯甲酸醋类防腐剂的定量方法[J].环境与健康杂志,1995,12(1):34-36.
[26]Soni M,Burdoek G A,Taylor L,et al.safety assessment of propylparaben:are view of the Published literature[J].Food and Chemical Toxieology,2001,39(6):513-532.
[27]Mahuzier P E,Altria K D,Cark B J.Selective and quantitative analysis of 4-hydroxybenzoate Preservatives by microemulsion electrokinetic chromatography[J].Journal of Chromatography A,2001,924(1-2):465-470.
[28]梁超,邓慧萍.水中内分泌干扰物质的研究现状及趋势城市给排水[J].2005 19(3):17-19.
[29]Kavelock R J.Research needs for risk assessment of health and environmental effects of endocrine disrupters:A report of the U.S.EPA-sponsored workshop[J].Environ Health Perspect,1996,104:715-740.
[30]何世华,梁增辉,晁福寰.环境雌激素测评方法研究进展[J].中国公共卫生,2002,18(10),1254-1256.
[31]Pesatori A C.Dioxin exposure and non-malignant health effects:a mortality study[J].Occup Environ Med,1998,55(2):126.
[32]Ankely G T,Johnson R D,Detenbeck N E,et al,Developmen of a research strategy for assessing the ecological risk of endocrine disruptors[J].Rev Toxicol,1997,18(3):231-267.
[33]Arcand-Hoy L D,Benson W H.Fish reproduction:An ecologi-cally relevant indicatior of endoerinc disruption[J].Environ Toxi-col Chem,1998,17(1):49-57.
[34] Ashby J, Tinwell H. Uterotrophicactivity of bisphenol A in the Immaturerat [J]. Environ Health Perspect, 1998, 106: 719-720.
[35] Randall Bolger. Rapid screening of environmental chemicals for estrgen receptor binding capacity [J].Environmental Health Perpectives, 1998, 106(9): 551-557.
[36] Kuiper G G, Enmark E, Pelto-Huikko M, et al. Cloning of a novel estrogen receptor expressed in rat prostate and ovary[J]. Proc Natl Sci USA, 1996, 93(3): 5925-5930.
[37] Didier G. Stable Luciferse Transfected Cells for Studing Receptor Biological Activity [J]. Biolumin Chemillumin, 1994, 9(3): 201-209.
[38] Zacharewski T. In vitro bioassays for assessing estrogenic substances. Environ Sci & Tech, 1997, 31:613-623.
[39] Junichi Ni, Koichi S. New Screening Methods for Chemicals with Coactivator [J]. Toxicology and Applied Pharmacology, 1999, 154(2): 76-83.
[40] Sumptre J P, Jobling S. Vitellogenesis as a biomarker for Estrogenic contamination of the qsuatic environment [J]. Environ Health Perspect, 1995, 103(3): 173-175.
[41] Palmer B D, Huth L K, Pieto D L, et al. Vitellogenin as abiomarker forxenobibtic estrogens in an amphibianmodel system [J]. Envrion Toxicol Chem, 1998, 17(1): 30-36.
[42] Monteverdi G H, Di Giulio R T. Anenzyme-linked imunosorbent assay for estrogenicity using primary hepatocyte cultures from the channel catfish (Ictalurus punctatus). Arch Environ Contam Toxicol, 1999,37: 62-69.
[43] Sherry J, Gamble A, Fielden M, et al. An ELISA for brown trout (Salmotrutta) vitellogenin and its use in bioassays for environmen-tal estrogens. Sci Total Environ, 1999, 225: 13-31.
[44] Chen C W, Hurd C, Vorojeikina D.P., et al. Trans criptional activation of the human estrogen receptor by DDT isomers and metabolites in yeast and MCF-7 cells. Biochem Pharmacol, 1997, 53: 1161-1172.
[45] Balaguer P., Joyeux A., Denisom M. S., et al. Assessing the estrogenic and dioxin-like activities of chemicals and complex mixtures using in vitror ecombinant receptor-reporter gene assays.Can J Physiol Pharmacol, 1996, 74: 216-222.
[46] Balaguer P., Francois F., Comunale F, et al. Reporter cell lines to study the etrogenic effects of xenoestrogens. Sci Total Environ, 1999, 233: 47-56.
[47] Legler J, vanden Brink C E, Brouwer A, et al. Development of astably transfected etrogen receptormediated luciferase reporter gene assay in the human T47D breast cancer cell line. Toxico Sci,1999,48:55-66.
[48] Pierrat B, Heery D M, Lemoine Y, et al. Function alanalysis of the Human estrogen receptor using aphenotypic transactivation in yeast. Gene, 1992, 119: 237-245.
[49] Rehmann K, Schramm K W, Kettrup A A. Applicability of a yeast oestrogen screen for the detection of oestrogen-like activities in environmental samples. Chemosphere, 1999, 38: 3303-3312.
[50] Vinggaard A M, Joergensen E C, Larsen J C. Rapid and sensitive reporter gene assays for detection of antiandrogenic and estrogenic effects of environmental chemicals. Toxical Appl Pharmacol, 1999. 155: 150-150.
[51]Han D,Tachibana H,Yamada K.Inhibition of environmental estrogen-induced proliferation of human breast carcinoma MCF-7cells by flavonoids[J].In Vitro Cell Dev Biol Anim,2001,37(5):275-282.
[52]Soto A M,Sonnenschein C,Chung K L,et al.The E-SCREEN assay as a tool to identify estrogens:An update on estrogenic environmental pollutants[J].In Estrogens in the Environment(Pro-ceedings from Estrogens in theEnvironment,Ⅲ:GlobalHealthImplications) Environ Health Perspect,1995,103(supp l7):113-122.
[53]Soto A M,Silvia R M,Sonnenschein C.A plasma-born specific inhibitor of the proliferation of human estrogen2sensitive tumor cancer cells(estrolycone-1)[J].J.Steroid Biochem Mol Boil,1992,43(7):703-712.
[54]Villalobos M,Olea N,Brotons J A,et al1.The E-screen assay:a comparison of different MCF-7 cell stocks[J].Environ Health Perspect,1995,103(9):844-850.
[55]Westley B.,Rochefort H.A secreted glycoprotein induced by estrogen in human breast cancer cell lines[J].Cell,1980,20(2):353-362.
[56]Schlumpf M,Cotton B,Conscience M,et al.In vitro and in vivo estrogenicity of UV screens[J].Environ Health Perspect,2001,109(3):239-244.
[57]石莹,张宏伟。环境内分泌干扰物的研究进展。国外医学卫生学分册,2006,33(6):342-347.
[58]Rogers K R,Williams L R.Biosensors for environmental monitoring:A regulatory perspective.TrAC,1995,14(7):289-294.
[59]翟俊辉,杨瑞馥.生物芯片、生物传感器和生物信息学.生物技术通报,2002,13(3):209-213.
[60]Marko Varga G,Emneus J,Gorton L,et al.Development of enzymebased amperometric sensors for the determination of phenolic compounds.TrAC,1995,14(7):319-328.
[61]韩梅梅,董国君,孙哲,等.生物传感器在环境监测中的应用.环境污染治理技术与设备2005,5(8):83-87.
[62]Suzuki H.Microfabrication of chemical sensors and biosensors for environmental monitoring.Materials Science and Engineering C:Biomimetic and Supermolecular Systems,2000,12(1):55-61.
[63]Ashley K.Developments in electrochemical sensors for occupational and environmental health applications[J].J.Hazard Mater,2003,102(1):1-10.
[64]Mestres V P.Chemosensitivity testing of human tumorsusing Si-sensor chips[J].Recent Results Cancer Res,2003(161):26-34.
[65]吴邦灿,费龙.现代环境监测技术[M].北京:中国环境科学出版社,1999.
[66]Frost M C,Meyerhoff M E.Implantable chemical sensors for real-time clinical monitoring:progress and challenges[J].Curr Opin Chem Biol,2002,6(5):633-642.
[67]Sigmundsson K,Masson G,Rice R,et al.Determination of active concentrations and association and dissociation rate constants of interacting biomolecules:an analytical solution to the theory for kinetic and mass transport limi-tations in biosensor technology and its experimental veri-fication[J].Biochemistry,2002,41(26):8263-8276
[68]Glark L C,Lyons C.Electrode systems for continuousmonitoring in car diovascular surger.Ann NY Acad Sci,1962,102:29-45.
[69]胡冠九,刘建琳,邹公伟.生物传感器在环境监测中的应用.环境监测管理与技术,1999,11(2):12-16.
[70]Karube I,Nomura Y,Arikawa Y.Biosensors for environmental control.TrAC,1995,14(7):295-299.
[71]Ricci F,Amine A,Palleschi G,et al.Prussian Blue based screen printed biosensors with improved characteristics of longterm lifetime and pH stability[J].Biosens Bioelectron,2003,18(2-3):165-174.
[72]Cheng H C,Abo M,Okubo A.Development of dimethyl sulfoxide biosensor using a mediator immobilized enzyme electrode[J].Analyst,2003,128(6):724-727.
[73]Ivanov A,Evtugyn G,Budnikov H,et al.Cholinesterase sensors based on screen-printed electrodes for detection of organophosphorus and carbamic pesticides[J].Anal BioanalChem,2003,377(4):624-631.
[74]Mitchell K M.Acetylcholine and choline amperometric enzyme sensors characterized in vitro and in vivo[J].Anal Chem,2004,76(4):1098-1106.
[75]Daniel R T,Klara T,et al.Electrochemical biosensors:recommended definitions and classification[J].Biosensors & Bioelectronics,2001,16(1-2):121-131.
[76]Gau J J,Esther H L,Bruce D,et al.AMEMS based amperometric detector for E.Coli bacteria using self-assembled monolayers[J].Biosensors & Bioelectronics,2001,16(9-12):745-755.
[77]Miquel A S,Arben M,Salvador A.Configurations used in the design of screen-printed enzymatic biosensor[J].A review,Sensors & Actuators B,2000,69(1-2):153-163.
[78]Tomas K,Petr S,A disposable amperometric immunosensor for 2,4-dichloro -phenoxyacctic acid[J].Analytic Chimica Acta,1995,304(3):361-368.
[79]Yulaev M F,Sitdikov R A.Dmitrieva N M,et al.Development of a potentiometric immunosensor for herbicide simazine and its application for food testing[J].Sensors and Actrators B,2001,75(1-2):129-135.
[80]Park S J,Taton T A,Mirkin C A.Array-based electrical detection of DNA with nanoparticle probes[J].Science.2002,295(22):1503-1506.
[81]Starodub N F,Dzatiev B B,Starodub V M.et al.Immunosensor for the determination of the herbicide simazine based on an ion-selective field-effect transistor[J].Analytica Chimica Acta,2000,424(1):37-43.
[82]Schuhmann W,Electron-transfer pathways in amperometric biosensor.Ferrocene- modified enzymes entrapped in conducting-polymer layers[J].Biosensors & Bioelectronics,1995,10(1-2):181 - 193.
[83]Ghindilis A L,Atanasov P,Wilkins M,et al.Immunosensors:electrochemical sensing and other engineering approaches[J].Biosensors & Bioelectronics,1998,13(1):113-131.
[84]Luppa P B,Sokoll L J,Chan D W.Immunosensors-principles and applications to clinical chemistry[J].Clinica Chimica Acta,2001,314(1-2):1-26.
[85]Keay R W,McNeil C J.Separation-free electrochemical immunosensor for rapid determination of atrazine[J].Biosensors & Bioelectronics,1998,13(9):963-970.
[86]Killard A J,Laura M.Amperometric separation-free immunosensor for real-time environmental monitoring[J].Analytica Chimica Acta 2001,427(2):173-180.
[87]Claycomb R W,Michael J.Delwiche.Biosensor for on-line measurement of bovine progesterone duringmilking[J].Biosensors & Bioelectronics,1998,13(11):1173-1180.
[88]Kreuzer M P,Pravda M,Ciara K.O,et al.Novel electrochemical immunosensor for seafood toxin analysis[J].Toxicon,2002,40(9):1267-1274.
[89]Faehnrich K A,Pravda M,Guilbault G G.Disposable amperometric immunosensor for the detection of polycyclic aromatic hydrocarbons(PAHs) using screen-printed electrodes[J].Biosensor and Bioelectronics,2003,18(1):73-82.
[90]Tahir Z M,Alcilja E C.A disposable membrane strip immunosensor[C].In:Proceedings of IEEESENSORS 2002,Orlando,FL,USA,2002,12-14.
[91]尹畅,刘健平,范静华.生物传感器在环境分析中的研究现状与前景.生物技术通报,2000(6):30-36.
[92]Marco M P,Gee S,Hammock B.D.Immunochemical techniques for environmental analysis:Ⅰ.Immunosensors.TrAC,1995,14(7):341-350.
[93]韩梅梅,董国君,孙哲,等.生物传感器在环境监测中的应用.环境污染治理技术与设备.2005,5(8):83-87.
[94]易志刚,熊雄.生物传感器在环境监测中的研究进展.工业安全与环保,2007,33(6):35-37.
[95]Wang J,Rivas G,Cai X,et al.DNA electrochemical biosensors for environmental moniitoring.A review.Anal Chim Acta,1997,347(1):1-8.
[96]Wang J,Chicharro M,Rivas G,et al.DNA biosensor for the detection of hydrazines.Anal Chem,1996,68(13):2251-2254.
[97]Nielson P,Egholm M,Berg R,et al.Sequence-selective recognition of DNA by strand displacement with a thymine substituted polyamide.Science,1991,254(8):1497-1500.
[98]万群,魏东芝,袁勤生,等.DNA芯片技术.生命的化学,1999,19(2):83-88.
[99]Maciejewska D,Szpakowska I,Wolska I,et al.DNA-based electrochemical biosensors for monitoring of bis-indoles as potential antitumoral agents,chemistry,X-ray crystallography.Bioelectrochemistry,2006,69(1):1-9.
[100]何冬梅,核肽酸-一种新型的反义物质.生命科学,1999,11(3):107-110.
[101]钟艳艳,王贵波,李宁,等.生物传感器现状与展望.重庆医学2005,34(5):659-662.
[102]Pandey P,Weetall H.Detection of aromatic compounds based on DNA intercalation using an evanescent wave biosensor.Anal Chem,1995,67(4):787-792.
[103]Wang J,Cai X,Johnson C,et al.Trace measurements of RNA by potentiometric stripping analysis at carbon paste electrodes.Anal Chem,1995,67(22):4065-4070.
[104]李宗义,邵强,郭伟云,等.用于环境监测的生物传感器.生物技术,2005,15(4):95-98.
[105]Zhang M,Li C.Fouling and natural organic matter removal in adsorbentm/membrane systems for drinking water treatment[J].Environ Sci Technol,2003,37(8):1663-1669.
[106]Shikha R,Anil K.Development and characterization of a novel immobilized microbial membrane for rapid determination of biochemical oxygen demand load in indusrial waste waters[J].Biosensors and Bioelectronics,2003,18:23-29.
[107]Shao C,Howe C,Porter A,et al.Novel cyanobacterial biosensor for detection of herbicides[J].Appl Environ Microbiol,2002,68(10):5026-5033.
[108]Mirasoli M,Feliciano J,Michelini E,et al.Internal response correction for fluorescent whole-cell biosensors[J].Anal Chem,2002,74(23):5948-5953.
[109]Christoph W,Marco C,Jan R.Measurement of biologically available naphthalene in gas and aqueous phases by use of a Pseudomonasputidabiosensor[J].Applied and Environmental Microbiology,2004,70(1):43-51.
[110]Michael N,Lars H,Mike S,et al.Bacterium-based NO-2biosensor for environmental applications[J].Applied and Environmental Microbiology,2004,70(11):6551-6558.
[111]Zhang M,Li C.Fouling and natural organic matter removal in adsorbentm/membrane systems for drinking water treatment[J].Environ Sci Technol,2003,37(8):1663-1669.
[112]鞠熀先,电分析化学与生物传感器[M].北京:科学出版社,2006.
[113]高体玉,冯军,慈云祥.细胞电化学的研究进展[J].化学进展,1998(10):305-311.
[114]Guo M L,Chen J H,Yun X B.Monitoring of cell growth and assessment of cytotoxicity using electrochemical impedance spectroscopy[J].Biochimica et Biophysica Acta(BBA) - General Subjects,2006,1763(3):432-439
[115]Li H N,Ci Y X,Feng J,et al.The voltammetric behavior of bone marrow of leukaemia and its clinical application[J].Bioeletrochem.Bioenerg.,1999,48(1):171-175.
[116]McConnell H M,Owicki J C,Parce J W,et al.The cytosensor microphysiometer:biological applications of silicon technology[J].Science,1992(257):1906-1912.
[117]Feng J,Ci Y X,Guo C M.et al.Voltammetric behavior of living cells T.shangaiensis and its bioanalytical application[J].Bioelectrochem.Bioenerg.1997(44):89-93.
[118]Li H N,Ci Y X.Electrochemical method for analyzing intracellular redox activity changes of the etoposide-induced apoptosis in HL-60 cells[J].Analytica Chimica Acta,2000,416(2):221-226.
[119]Yang L.Li Y,Griffis C L,et al.Interdigitated microelectrode(IME) impedance sensor for the detection of viable Salmonella typhimurium[J].Biosens.Bioelectron.,2004,19(10):139-1147.
[120]Dheilly A.Linossier I,Darchen A,et al.Monitoring of microbial adhesion and biofilm growth using electrochemical impedancemetry[J].Appl.Microbiol.Biotechnol.2008,79(1):57-164.
[121]Zhu L L,Gao N,Zhang x l,et al.Accurately measuring respiratory activity of single living cells by scanning electrochemical microscopy[J].Talanta,2008.77(2):804-808.
[122]Chen H,Heng C K,Puiu P D,et al.Detection of Saccharomyces cerevisiae immobilized on self-assembled monolayer(SAM) of alkanethiolate using electrochemical impedance spectroscopy[J].Anal.Chim..Acta,2005,554(1-2):52-59.
[123]Fojta M.Fojtova Mi,Havran L,et al.Electrochemical monitoring of phytochelatin a ccumulation in Nicotiana tabacum cells exposed to sub-cytotoxic and cytotoxic levels of cadmium [J]. Analytica Chimica Acta, Volume, 2006, 558(1-2): 171-178.
[124] Diakowski P M, Ding Z F. Interrogation of living cells using alternating current scanning electrochemical microscopy (AC-SECM) [J]. Chem. Chem. Phys., 2007(9): 5966 -5974.
[125] Xiao C, Lachance B, Sunahara G, et al. Assessment of Cytotoxicity Using Electric Cell-Substrate Impedance Sensing: Concentration and Time Response Function ApproachAnal [J]. Chem., 2002,74(22): 5748 -5753.
[126] Ding L, Du D, Wu J, et al. A disposable impedance sensor for electrochemical study and monitoring of adhesion and proliferation of K562 leukaemia cells [J]. Electrochem. Commun., 2007,9(5): 953-958.
[127] Fojta M, Fojtova M, Havran L, ET AL. Construction and use of an integrated electrochemical device for the detection of biologically relevant compounds released from non-adherent cells: Application for the electrochemical determination of nitric oxide produced by human U937 cells [J]. Analytica Chimica Acta,2006, 558 (1-2): 171-178.
[128] Arndt S, Seebach J, Psathaki K, et al. Bioelectrical impedance assay to monitor changes in cell shape during apoptosis [J]. Biosens. Bioelectron. ,2004,19(26): 583-594.
[129] Woolley D E, Tetlow L C, Adlam D J, et al. Electrochemical Monitoring of Anticancer Compounds on the Human Ovarian Carcinoma Cell Line A2780 and Its Adriamycin- and Cisplatin-Resistant Variants[J]. Exp. Cell Res., 2002, 273(1): 65-72.
[130] Li H N, Ci Y X, Feng J, et al. The voltammetric behavior of bone marrow of leukaemia and its clinical application [J]. Bioelectrochem. Bioenerg., 1999,48(1): 171-175.
[131] Chen K, Chen J H, Guo M L, et al. Electrochemical Behavior of MCF-7 Cells on Carbon Nanotube Modified Electrode and Application in Evaluating the Effect of 5-Fluorouracil [J]. Electroanalysis.2006, 18(12): 1179-1185.
[132] Cheng W, Ding L, Lei J P, Effective Cell Capture with Tetrapeptide-Functionalized Carbon Nanotubes and Dual Signal Amplification for Cytosensing [J]. Anal. Chem..2008(80): 3867-872.
[133] Chen J, Du D, Yan F, et al. Electrochemical Antitumor Drug Sensitivity Test for Leukemia K.562 Cells at a Carbon-Nanotube-Modified Electrode [J]. Chem. Eur. J., 2005(11): 1467-1472.
[134] Yan F, Chen J, Ju H X. Immobilization and electrochemical behavior of gold nanoparticled leukemia K562 cells and application in drug sensitivity test [J]. Electrochem. Commun. 2007, 9(2): 293-298.
[135] Dua D, Liu S L, Chen J, et al. Colloidal gold nanoparticle modified carbon paste interface for studies of tumor cell adhesion and viability [J]. Biomaterials 2005, 26(33): 6487-6495.
[136] Ding L, Hao C, Xue Y D, et al. A Bio-Inspired Support of Gold Nanoparticles-Chitosan Nanocomposites Gel for Immobilization and Electrochemical Study of K562 Leukemia Cells [J].Biomacromolecules, 2007, 8(4): 1341-1346.
[137] Feng J, Luo G A, Jian H Y, et al. Voltammetric Behavior of Tumor Cells U937 and Its Usefulness in Evaluating the Effect of Caffeic Acid [J]. Electroanalysis, 2000,12(7): 513-516.
[138] Li J, Liu X, Guo M, et al. Electrochemical Study of Breast Cancer Cells MCF-7 and Its Application in Evaluating the Effect of Diosgenin[J]. Anal. Sci. 2005, 21(5): 561-564.
[139] Sucheta A, Ackrell B A C, Cochran B, et al. Diode-like behaviour of a mitochondrial electron-transport enzyme [J]. Nature 1992(356): 361-362.
[140] Woolley D E, Tetlow L C, Adlam D J, et al. Electrochemical Monitoring of Anticancer Compounds on the Human Ovarian Carcinoma Cell Line A2780 and Its Adriamycin- and Cisplatin-Resistant Variants[J]. Exp. Cell Res. 2002, 273(1): 65-72.
[141] Prabhulkara S, Alwarappana S, Liu G D. Amperometric micro-immunosensor for the detection of tumor biomarker [J]. Biosensors and Bioelectronics 2009, 24(12): 3524-3530.
[142] Ci Y X, Zhai Q, Wang S, et al. Voltammetric studies of the effect of Cisplatin-liposome on Hela cells [J]. Talanta. 2001, 55(4): 693-698.
[143] Feng J, Ci Y X, Lou J L, et al. Voltammetric behavior of mammalian tumor cells and bioanalytical applications in cell metabolism [J]. Bioeletrochem. Bioenerg., 1999, 48(1): 217-222.
[144] Li H N, Ci Y X, Feng J, et al. The voltammetric behavior of bone marrow of leukaemia and its clinical application [J]. Bioeletrochem. Bioenerg., 1999,48(1): 171-175.
[145] Ci Y X, Feng Z J, Jiang W, et al. The voltammetric behavior of Saccharomyces cerevisiae[J].Bioeletrochem. Bioenerg., 1997,43(2): 293-296.
[146] Harrison R. structure and function of xanthine ocidoreductase: where are we now [J]. Free Rad. Biol.Med., 2002, 33(6): 774 - 797.
[147] Xiao F. Ruan C P, Li J G. et al. Voltammetric Determination of Xanthine with a Single-Walled Carbon Nanotube-lonic Liquid Paste Modified Glassy Carbon Electrode [J]. Electroanalysis[J]. 2008. 20(4):361-366.
[148] 程介克.单细胞分析[M].北京:科学出版社, 2005,8.
[149] F. He, Q. Shen, H. Jiang, J. Zhou, J. Cheng, D. D. Guo, Q. N. Li. X. M. Wang, D. G Fu, B. A. Chen,Rapid identification and high sensitive detection of cancer cells on the gold nanoparticle interface by combined contact angle and electrochemical measurements [J]. Talanta, 2009(77): 1009 - 1014.
[150] Andreescu S. Sadik O A, McGee D W. Effect of natural and synthetic estrogens on A549 lung cancer cells: correlation of chemical structures with cytotoxic effects[J]. Anal. Chem. 2004(76): 2321-2330.
[151] L.J. Yang, Y.B. Li, GF. Erf, Interdigitated array microelectrode-based electrochemical impedance immunosensor for detection of Escherichia coli O157:H7 [J]. Anal. Chem., 2004(76): 1107-1113.
[152] H. Chen, C. K. Heng, P. D. Puiu, X. D. Zhou, A. C. Lee, T. M. Lim, S. N. Tan. Detection of Saccharomyces cerevisiae immobilized on self-assembled monolayer (SAM) of alkanethiolate using electrochemical impedance spectroscopy [J]. Anal. Chim. Acta, 2005(554): 52-59.
[153] Y. Torisawa, T. Kaya, Y. Takii, D. Oyamatsu, M. Nishizawa, T. Matsue, Scanning Electrochemical microscopy-based drug sensitivity test for a cell culture integrated in silicon microstructures scanning electrochemical microscopybased drug sensitivity test for a cell culture integrated in silicon microstructures [J]. Anal. Chem., 2003(75): 2154-2158.
[154] Xiao C. Lachance B, Sunahara G, Luong J H T. Assessment of cytotoxicity using electric cell substrate impedance sensing:concentration and time response function approach[J],Anal.Chem.,2002(74):5748-5753.
[155]Cheng W,Ding L,Lei P J,et al.Effective cell capture with tetrapeptide-functionalized carbon nanotubes and dual signal amplification for cytosensing and evaluation of cell surface carbohydrate[J].Anal.Chem.,2008(80):3867-3872.
[156]Wu D M,Fu G L,Fang H Z,et al.Studies on the origin of the voltammetric response of the PC-3 cell suspension[J].Talanta,2009(78):602-607.
[157]石莹,张宏伟。环境内分泌干扰物的研究进展。国外医学卫生学分册,2006,33(6):342-347.
[158]Rotchell J,Ostrander G..Molecular markers of endocrine disruption in aquatic organisms[J].Toxicological Environmental Health,Part B,2003(6):453-495.
[159]张静云,吕剑,何义亮等.环境雌激素的生物检测与应用[J].环境科学与技术,2007,30(2):100-102.
[160]Larkin P,Folmar L C,Hemmer M J,et al.Expression profiling of estrogenic compounds using a sheepshead minnow cDNA macroarray[J].Environmental Health Perspective,2003(111):839-846.
[161]Wang D,McKague B,Liss S,et al.Gene expression profiles for detecting and distinguishing potential endocrine-disrupting compounds in environmental samples[J].Environmental Science and Technology,2004,38(23):6396-6406.
[162]F.Jun,Y.X.Ci,J.L.Lou,X..Q.Zhang.Voltammetric behavior of mammalian tumor cells and bioanalytical applications in cell metabolism.Bioelectrochem.Bioenerg[J].1998(45):247-222.