摘要
蓖麻毒素(Ricin Toxion, RT)是一种核糖体失活蛋白,由A、B链组成,小鼠LD50仅为7-10μg/kg。目前国内外对蓖麻毒素的研究多为用于抗肿瘤的免疫毒素的制备以及中毒机理的研究,由于蓖麻毒素的毒性高,从摄入到死亡仅需1~2h,致死量极小,感官无法辨认,蓖麻毒素类毒素和抗毒素正在研究中,目前尚无用于人的特效药,因此,蓖麻毒素的快速检测,特别是在被人体摄入之前就能够高效灵敏的检测到,就显的极为重要。
本试验将制备的抗蓖麻毒素腹水单抗纯化后进行抗原表位的分析,结果显示4D8、4D12、1H4三株单抗的抗原决定簇均位于蓖麻毒素A链上。对4D8标记了辣根过氧化物酶和FITC,基于表面等离子共振仪与流式细胞仪分析后,分别建立了夹心ELISA、IMS-ELISA、SPR与液态芯片检测蓖麻毒素的方法。
1.夹心ELISA检测蓖麻毒素方法的最佳条件为:45ug/ml抗RT的多克隆抗体为捕获抗体、1:4000倍稀释的HRP-4D8为检测抗体、5%脱脂乳+2%PEG20000为封闭剂,洗去检测抗体后用TMB在37℃下显色20min,测定OD_(450);该方法不跟与蓖麻毒素结构相近的其它几种毒素发生交叉反应,最低检出浓度为5ng/ml,线性范围为0.1-1ug/ml,对纯净水的回收率为107.6%。
2. IMS-ELISA检测蓖麻毒素方法的最佳条件为:将多抗与磁珠偶连并封闭后用PBST缓冲液悬浮在EP管中,加入抗原后37℃轻微振荡孵育1h,每15min吹打重悬一次,清洗后加入4000倍稀释的HRP-4D8检测抗体200μl孵育,清洗后加TMB底物在37℃下显色20min,测定OD_(450);该方法不跟与蓖麻毒素结构相近的其它几种毒素发生交叉反应,最低检出浓度为50ng/ml,当RT浓度介于0.2-10ug/ml之间时,呈现极好的线性关系,对纯净水、秋梨膏、纯牛奶的回收率分别为95%、93%、68%。
3.由SPR动力学分析了四株抗蓖麻毒素单抗的亲和性:并建立了SPR检测蓖麻毒素与相思子毒素的SPR检测方法,对蓖麻毒素的线性范围为0-2000ng/ml之间,最低检出浓度为0.05ng/ml,对相思子毒素的线性范围为250ng/ml-2000ng/ml,最低检出浓度为0.5ng/ml。对两种毒素的混合物同时检测时与单独检测的效果一样。
4.本研究同时基于流式细胞仪与微球载体建立了对蓖麻毒素的检测方法,对蓖麻毒素的线性范围为10-1000ng/ml之间,最低检出浓度小于0.1ng/ml。
5.采用PCR法扩增出蓖麻毒素B链基因,并将其与pMD18-T克隆载体相连接,经序列分析正确后插入到表达载体pET-28a中,构建重组表达质粒pET-28a-RTB,将构建好的重组质粒转化入BL21感受态细胞中,经IPTG于37℃诱导表达后,得到的融合表达的目的蛋白,经SDS-PAGE分析,相对分子量约为31KD,利用Western bloot与间接ELISA鉴定证明其具有反应原性,为制备RTB单克隆抗体及提高检测方法的敏感性打下基础。
This paper reported that 4D8、4D12 and 1H4 are McAb anti-ricin, which purificated by protein-G chromatographic column, and ELISA data showed that their antigenic determinant were located on ricin A china. we labered 4D8 with horse radish peroxidase(HRP) and FITC respectively, then established ELISA、IMS-ELISA、surface plasma resonance(SPR) to detect ricin. test data indicated:
1.The optimum condition of sandwich ELISA assays for ricin is: diluted RT polyclonal antibody to 45 ug/ml as capture Ab, diluted HRP-4D8 to 4000 times as detection Ab, chose 5%skimmed milk+2%PEG20000 as blockage, add TMB substrate solution after wash away detection Ab, incubation at 37℃for 20min. read OD_(450). This method don’t have cross-reaction with other resemble toxion, minimum detectable concentration is 5ng/ml, The detection range is 0.1-1ug/ml, standard recovery rate to purified water is 107.6%.
2.The optimum condition of IMS-ELISA assays of ricin is: coupling beads with polyclonal antibody anti-ricin, add PBST in EP tube to diluted beads after block, add assay sample incubate for 1h at 37℃with slow tilt rotation. Resuspension with micropipettor every 15min, diluted HRP-4D8 to 4000 times, add 200μl as detection Ab, add TMB substrate solution 100μl after wash away detection Ab, incubat at 37℃for 20min. read OD_(450). This method don’t have cross-reaction with other resemble toxion, minimum detectable concentration is 50ng/ml, The detection range is 0.2-10ug/ml, standard recovery rate to purified water、pear jam and milk respectively is 95%、93%、68%.
3.Kinetic analysis base on SPR result showed the affinity to ricin:4D8 KA=3.16×10~(12) ,KD=3.16×10~(-12);4D12 KA=1.53×10~(13), KD=6.55×10~(-14);2F KA=4.1×10~8, KD=2.44×10~(-9);1C KA=1.86×10~(10), KD=5.38×10~(-11). Establish the metherd base on SPR assay for ricin and abrin, The detection range for ricin is 0-2000ng/ml, for abrin is 250-2000ng/ml; minimum detectable concentration for ricin is 0.05ng/ml, for abrin is 0.5ng/ml; And the effect of detect single toxion is equal to mixture of ricin and abrin.
4. Establish the metherd base on FCM and beads assay for ricin. The detection range for ricin is 10-1000ng/ml, minimum detectable concentration for ricin is 0.1ng/ml.
5. The DNA of RTB was cloned from the ricin DNA by PCR and inserted into the carrier pMD18-T, The fusion expression vector RTB-pET28 was contructed and transformed to E.coli BL21 after sequenced wand. The expressed fusion protein after induction with IPTG at 37℃. SDS-PAGE and Western blotting analysis showed that the 31KD fusion protein was expressed correctly in E.coli. and the bioactivity was detected by indirect ELISA. The results provided reliable basis for the specificity of ELISA detect ricin.
引文
[1] 董巨莹等. 蓖麻毒素及其修饰物对U937细胞IL21B和TNF2A的诱导作用. [J] 细胞与分子免疫学杂志,1999,15: 123-125
[2] 贺永怀等. 一种快速简便的提取和纯化蓖麻毒素的方法. [J] 生物化学与生物物理进展,1987,6: 64-66
[3] 郑成,雷德柱,雷雨. 蓖麻毒素提取. [J] 广东化工,2003,5: 1-4
[4] 陈志慧.蓖麻毒素的研究与应用进展. [J] 化学世界,5: 309-312
[5] Sandvig K, van Deurs B.Transport of protein toxins into cells:pathways use dcholera toxin and Shiga toxin.FEBS Lett.2002,529(1): 49-53
[6] Monfort W, Villafranca JE, Monzingo AF, et al. The three dimensional structure of ricin at 2.8?.J Biol Chem 1987; 262: 5398-5403
[7] Rutenber E, Robertus JD. Structure of ricin b-chain at 2.5? resolution. Proteins, 1991, 10: 260-269
[8] Monzingo AF, Robertus JD: X-ray analysis of substrate analogs in the ricin a-chain active site. J Mol Biol, 1992, 227: 1136-1145
[9] Lord JM, Roberts LM, Robertus JD. Ricin: structure, mode of action, and some current applications. FASEB [J], 1994, 8: 201-208
[10] Weston SA, Tucker AD, Thatcher DR, Derbyshire DJ, Pauptit RA.X-ray structure of recombinant ricin a-chain at 1.8 ?. resolution. J Mol Biol, 1994, 266: 1043-1049
[11] Day PJ, Ernst SR, Frankel AE, et al. Structure and activity of an active substitution of ricin a chain. Biochemistry 1996, 35: 11098-11103
[12] Wales R, Richardson PT, Roberts LM; et al. Mutational analysis of the galactose binding ability of recombinant ricin b chain. J Biol Chem, 1991, 266: 19172-19179.
[13] Olsnes S., Pihl A. in "Molecular action of toxins and viruses (Cohen P, Van Heyningen S.eds )Elsevier Biomedical Press, Amsterdam-New York-Oxford. 1982: 51-105.
[14] 孙嗣梅.蓖麻毒素A链及其突变体的表达、纯化和分析.学位论文 2005
[15] Barbieri L,Battelli MG,Stirpe F.Ribosome-inactiviting proteins from plants[J]. Biochim.Biophys. Acta,1993,1154: 237-282
[16] Olson MA.Ricin A chain structural determinant for binding substrates analogues: a molecular dynamics simulation analysis[J]. Proteins,l997,27: 80-95
[17] Day PJ, Ernst SR,Frankel AE, et al. Structure and activity of an active site substitution of ricin A chain[J]. Biochemistry,1996,35: 11098-11103
[18] Marsden CJ, Smith DC, Roberts LM, et al. Ricin: current understanding and prospects for an antiricin vaccine[J]. Expert Rev. Vaccines, 2005, 4(2): 229-237
[19] Yan X, Hollis T, Svinth M, et al.Structure-based identification of a ricin inhibitor[J]. Mol .Biol,1997,266: 1043-1049
[20] 马嘉惠.蓖麻毒素的研究与应用[J]. 氨基酸和生物资源,1999,21(2): 56-58
[21] Ru Tenber E, Robertus J. D. Structure of ricin B-chain at 2.5A resolution[J]. Proteins, 1991, 10(3): 260～269
[22] 赵青余,桂荣,那日苏. 蓖麻饼脱毒的研究进展[J ] .饲料工业,2002 ,23 (11) : 35 -39
[23] 葛利萍,詹金彪.蓖麻毒素在细胞内的运输和转位途径研究进展[J].浙江大学学报(医学版)2003,32(4): 542-546
[24] Olsnes S, Fernandez-Puentes C, Carrasco L, et al. Ribosome inactivation by the toxic lectins abrin and ricin. Kinetics of the enzymic activity of the toxin A-chain. Eur.J.Biochem, 1975, 60: 281-288.
[25] Balint GA. Ricin: the toxic protein of castor oil seeds. Toxicology. 1974, 2(1): 77-102.
[26] Wannemaker R. Army Medical Research Institute of Infectious Diseases, Fort Detrick,Frederick, Md. Personal communication, September 1994.
[27] Ranz DR, Jaax NK. Medical aspects of chemical and biological warfare, Chapter 32 Ricin toxin, Falls Church, VA: Office of the Surgeon General. 1997, :631-642.
[28] POISINDEX?. Editorial Staff: Castor beans (Management/Treatment Protocol). In:Klasco RK,ed. POISINDEX ? System. Greenwood Village; Colorado:Thomson MICROMEDEX. (Edition expires [Nov, 2003].
[29] Olsnes S, Refsnes K, Pihl A. Mechanism of action of the toxic lectins abrin and ricin. Nature, 1974a, 249: 627-631.
[30] Olsnes S, Saltvedt E, Pihl A. Isolation and comparison of galactose-binding lectins from Abrus precatorius and Ricinus communis. J.Biol.Chem, 1974b, 249: 803-810.
[32] Fu T, Burbage C, Tagge Ep, et al. Ricin toxin contains three lectin sites which contribute to its in vivo toxicity. Int J Immunopharmacol, 1996, 18: 685-692
[33] 赵英.蓖麻毒素毒性作用机理研究进展[J]. 中华预防医学杂志 1999,1,33: 54-55
[34] Endo Y, Tsurugi K. The characterics of the enzymatic activity of ricin A-chain with ribosome and with rRNA [J]. Biol Chem. 1988; 263(18): 8735-8739
[35] 赵丹.蓖麻毒素的提取及杀虫效果研究.硕士学位论文.2005
[36] Rigerio L, Vitale A, Lord J M ,et al. Free ricin A chain proricin and native toxin have different cellular fates when express in tobacco proplasts.[J]. Bio Chem.1998;273(23): 14194-14199
[37] Tracey KJ , Beutler B, Lowry SF, et al. Shock and tissue injury induced by recombinant human cachetin. Science, 234: 470-474
[38] Licastro F, Morini MC, Bolognesi A , et al. Ricin induced the production of tumor necrosis factoralpha and interleukin-1 beta by human peripheral-blood mononuclear cells. Biochem J , 1993,294 (Pt2) : 517-520
[39] Muldoon DF, BagchiD, Hassoun EA , et al. Themodulatingeffects of tumor necrosis factor alpha antibody on ricin-inducedox-idative stress in mice. J Biochem Toxico l, 1994, 9: 311-318
[40] 董巨莹,王剑波,王文学. 蓖麻毒素诱导小鼠肝脏产生肿瘤坏死因子[J]. 第四军医大学学报, 1997,18: 78
[41] Muldoon DF, Stohes SJ. Ricin-induced oxidative stress in mice . Toxicologist, 1991, 11: 214
[42] Muldoon DF, Hassoun EA , Stohs SJ. Ricin-induced hepatic lipid peroxidation, glutathione depletion and DNA single-strand break s in mice. Toxicon, 1992, 30: 977-984
[43] Corwin AH. Toxic constituents of the castor bean. JM ed PharmChem, 1961, 4: 483-496
[44] Muldoon DF, Hassoun EA , Stohes SJ. Role of iron in ricin-in-duced lipid peroxidation and superoxide production. Res CommunMol Pathol Pharmacol, 1996, 92: 107-118
[45] Leek MD, Griffiths GD, Green MA . Intestinal pathology following intramuscular ricin poisoning. J Pathol, 1989, 159: 329-334
[46] Waring P. DNA fragmentation induced in macrophages by gliotoxin does not require protein synthesis and is preceded raised inositol triphosphate levels. J Bio Chem, 1990, 265: 14476-14480
[47] Khan T, Waring P. Macrophage adherence prevents apoptosis induced by ricin. Eur J Cell Bio , 1993, 62: 4062-4141
[48] 江渝.程序型细胞死亡的生物化学基础.国外医学临床生物化学与检验学分册, 1996, 17: 178-182
[49] Jennifer Audi MD,Martin Belson MD et al. Ricin Poisoning A Comprehensive Review. CLINICAL REVIEW: 2342-2351
[50] Kirsten Sandvig,Bovan Deurs. Endocytosis and intracellular sorting of ricin and Shigatoxin. FEBS Lett, 1994:346: 99
[51] S Ramak rishnan, D Fryxell,MohanrajD, et al. Cytotoxic conjugates containing translational inhibitory proteins. A nn Rev Pharmaco Toxico, 1992; 32: 579
[52] Riccobono Fabio, FianiML. Mannose receptor dependent up take of ricin A1 and A2 chains by macrophage.Carbohydr Res, 1996; 282 (2) : 285
[53] 裴武红.蓖麻毒素内化研究进展[J]. 免疫学杂志1997,13,4: 275-277
[54] Van Deurs B, Petersen OW, Olsnes S, et al. The ways of endocytosis. Int. Rev Cytol,1989, 117: 131-177.
[55] Doan LG. Ricin: mechanism of toxicity, clinical manifestations ,and vaccine development.J Toxico-Clin Toxico, 2004, 42(2): 201-208.
[56] Van Deurs B, Sandvig K. Transport of protein toxins into cells: pathways used by ricin,cholera toxin and shiga toxin. FEBS Lett, 2002, 529: 49-53.
[57] Sphyris N, Lord JM, Wales R, et al. Mutational analysis of the ricinus lectin b-chains.Galactose-binding ability of the gamma subdomain of riciuus communis agglutinin b-chain. J Biol Chem, 1995, 270: 20292-20297.
[58] Roberts L, Wales R, Lard JM. Addtion of an endoplasmic reticulum retrieval sequence to ricin A chain significantly increase its cytotoxicity to mammalian cells. J Biol Chem. 1993
[59] Wales R, Chaddock JA, Roberts LM, et al. Addition of ER retention signal to ricin A chain increase the cytotoxicity of the holotoxin. Exp Cell Res.1992,203
[60] Bilge A, Clark JH. Degradation of ricin A chain by endosomal and lysosomal enzyme: the protective role of ricin B chain. Ther Immunol.1994; 1:197
[61] 董巨莹.蓖麻毒素的毒作用及其信号转导的分子机理研究. 1999
[62] Sandvig K, Grimmer S, Lversen TG, et al.Ricin transport into cells:studies of endocytosis and intracellular transport[J]. Intel J Med Microbiol, 2000,290(4-5): 415-420
[63] Vogel U,Sandvig K, Van Deurs B. Expression of caveolin-1 and polarized formation of invaginated caveolae in Caco-2 and MDCKIIcells[J].J Cell Sci,1998,111: 825-832
[64] Sandvig K and Van Deurs B. Entry of ricin and Shiga toxin into cells: molecular mechanisms and medica perspectives[J]. EMBOJ, 2000, 19 (22):5943-5950
[65] Sandvig K and Van Deurs B. Endocytosis and intracellular transport of ricin : recent discoveries[J].FEBS Letters, 1999, 452 (1-2): 67-70
[66] Garred?,Rodal SK, Van Deurs B, et al. Reconstitution of clathrin-independent endocytosis at the apical domain of permeabilized MDCK II cells :requirement for a Rho-family GTPase [J].Traffic, 2001, 2: 26-36
[67] Llorente A, Van Deurs B, Garred ?,et al. Apical endocytosis of ricin in MDCK cells is regulated by the cyclooxygenase pathway [J].J Cell Sci,2000,113(pt7): 1213-1221
[68] Rodal SK,Skretting G, Garred ?,et al. Extraction of cholesterol with methyl-beta- cyclodextrin perturbs formation of clathrin一coated endocytic resicles [J].Mol Biol Cell,1999,10(4): 961-974
[69] Subtil A, Gaidarov I, Kobylarz K, et al. Acute cholesterol depletion inhibits clathrin-coated pits budding[J].Proc. Natl. Acad. Sci.USA, 1999, 6 (12): 6775 -6780
[70] Lorente A, Prydz K, Sprangers M, et al .Proteoglycan synthesis is increased in cells with impaired clathrin-dependent endocytosis[J]. J Cell Sci,2001,114: 335-343
[71] 葛利萍,詹金彪:蓖麻毒素在细胞内的运输和转位途径研究进展[[J]. 浙江大学学报(医学版)2003,32 ( 4 ) : 542-546
[72] Edward Tagge, John Chandler, Bor Luen Tang. Cytotoxicity of KDEL-terminated ricin toxins correlates with distribution of the KDEL receptor in the Golgi. J Histochem Cytochen. 1996;44(2):159
[73] Iversen TC,Skretting G, Llorente A, et al. Endosome to Golgi transport of ricin is independent of clathrin and of the Rab-9 and Rab-11 GTPase[J].Mol Biol Cell, 2001,12:2099-2107
[74] Ghosh RN, Mallet WG, Soe TT, et al. An endocytosed TGN38 chimeric protein is delivered to the TGN after traffcking through the endocytic recycling compartrnent in CHO cell [J]. J Cell Biol,1998,142(4):923-936
[75] Grimmer S, Iversen TG, Van Deurs B, et al .Endosoroe to Golgi transport of ricin is regulated by cholesterol[J].Mol Biol Cell, 2000, 11(12):4205-4216
[76] Simpson JC, Lord JM, Roberts LM. Point mutantions in the hydrophobic C- terminal region of ricin Aindicate that Pro250 plays a key role in the membrane translocation. Eur J Biochem. 1995;232:458
[77] Roberts LM, Simpson JC, Lord JM. Catalysic and cytotoxic activities of recombinant ricin A chain mutant with charged residues added at the carboxyl terminus protein. Expression Purif. 1995;6:665
[78] Michalak M, Corbett EF, Mesaeli N, et al. Calreticulin :one protein, one gene, many functions[J].Biochem J, 1999, 344 (pt2):281-292
[79] Day PJ, Owens SR, Wesche J, et al. An interaction between ricin and calreticulin that may have implications for toxin trafficking[J].J Biol Chem, 2001, 276(10):7202-7208
[80] Pagny S, Cabanes-Macheteau M, GillikinJW,et al .Protein recycling from the Golgi apparatus to the endoplasmic reticulum in plants and its minor contribution to calreticulin retention[J].Plant Cell, 2000,12 (5):739-756
[81] Simpson JC, Roberts LM, Romisch K, et al .Ricin A chain utilises the endoplasmic reticulum-associated protein degradation pathway to enter the cytosol of yeast [J].FEBS Letters, 1999, 459(1):80-84
[82] Franco Dosio, Antonia Franceschi, Maurizio Ceruti, et al .Enhancement of ricin toxin A chain immunotoxin activity: synthesis, ionophoretic ability and in vitro activity of monensin derivatives. Biochem Pharmacol. 1996;52(1):157
[83] 裴武红.蓖麻毒素内化研究进展 [J].免疫学杂志 1997,13,4,275:277
[84] Wesche J, Rapak A, Olsnes S. Dependence of ricin toxicity on translocation of the toxin A-chain from the endoplasmic reticulum to the cytosol [J]. J Biol Chem,1999,274(48): 34443-34449
[85] Argent RH, Parrott AM, Day PJ, et al. Ribosome-mediated folding of partially unfolded ricin A chain[J]. J Biol Chem, 2000, 275(13): 9263-9269
[86] 赵丹.蓖麻毒素的提取及杀虫效果研究. 硕士学位论文. 2005
[87] Maruniak J E, Fiesler S E, Mcguire P M .Susceptibility of Insect Cells and Ribosomes to Ricin[J].Comparative Biochemistry and Physiology,1990, 96: 543-548
[88] 赵善欢.昆虫毒理学[M].北京,中国农业出版社,1999
[89] 赵善欢.植物化学保护[M].北京:中国农业出版社,2002
[90] 丁锦华,苏建亚. 农业昆虫学(南方本)[M]. 北京:中国农业出社,2002
[91] 何耀安. 蓖麻毒素的提取分离及对青藏高原鼠的防治研究 2006
[92] 詹金彪,周佩衍. 抗直肠癌免疫HCMcAb83 :Ricin 的制备及其特异杀伤作用[J ]. 中国免疫学杂志,1991 ,5 (3) :188～189.
[93] Engert a , Vitetta E. A phase I study of anti-CD25 ricin A chain immunotoxin in patients with refractory Hodgkin′s Lymphorma[J ] . Blood ,1997 ,89(2): 403～410.
[94] Johson BE , Kelley MJ . Autocrine growth factors and neuroendocrine markers in the development of small cell lung cancer[J ] . Oncology Huntint ,1998 ,12: 11～14.
[95] Gelber BE , Vitetta ES. Effect of immunosuppressive agents on the immunogenicity and efficacy of an immunotoxin in mice[J ] . Clin Cancer Res ,1998 ,4(5):1297～1304.
[96] Francisco JA , Siegall CB. In vivo efficacy and toxicity of asingle-chain immunotoxin targeted to CD40[J ] . Blood ,1997 ,89 (12) :4493～4450.
[97] Alamatsu Y, Murphy JC , Leung SO. A single-chain immunotoxin against carcinoembryonic antigen that suppresses growth of colorectal carcinoma cells [ J ] . Clin Cancer Res ,1998 ,4 (11) :2825～2832.
[98] 邹立波.蓖麻毒素与肿瘤治疗[J]. 国外医学临床生物化学与检验学分册.2001,22,6: 290-292
[99] 陈志慧.蓖麻毒素的研究与应用进展[J]. 化学世界,2004,5: 309-313
[100] 康凤龙.升麻善解蓖麻籽中毒[J].国医论坛 2005,7,20
[101] Yan C, Rill WL, Malli R, et al. Dependence of ricin toxoid vaccine efficacy on the structure of poly (lactide-co-glycolide) microparticle carriers. Vaccine. 1995, 13: 645.
[102] Griffiths GD, Phillips GJ, Bailey SC. Comparison of the uality of protection elicited by toxoid and peptide liposomal vaccine formulations against ricin as assessed by markers of inflammation. Vaccine. 1999, 17: 2562.
[103] Smallshaw JE, Firan A, Fulmer JR, et al. A novel recombinant vaccine which porotects mice against ricin intoxication. Vaccine. 2002, 20: 3422.
[104] Clarke E. G. The detection of ricin. J. Pharm. Pharmacol. 1953, 5: 458-459.
[105] Poli M. A;Rivera V. R.; Hewetson J. F; et al. Detection of ricin by colorimetric and chemiluminescence ELISA. Toxicon 1994, 32: 1371-1377.
[106] Shyu H. F.; Chiao D. J.; Liu H. W.; et al. Monoclonal antibody-based enzyme immunoassay for detection of ricin. Hybrid. Hybridomics 2002, 21: 69-73.
[107] Boroda E.; Jaynes W. F.; Zartman R. E. et al. Enzyme-Linked ImmunoSorbent Assay measurement of castor toxin in soils. Communications In Soil Science and Planr Analysis 2004, 35:1185-1195.
[108] Delehanty J. B.; Ligler F. S. A microarray immunoassay for simultaneous detection of proteins and bacteria. Anal. Chem. 2002, 74 (21): 5681-5687. [110] Poli MA, Revera VR, Hewetson JF, et al. Detection of ricin by colorimetric and chemiluminescence ELISA. Toxicon. 1994, 32: 1371.
[109] Shyu RH, Shyu HF, Liu FW, et al. Colloidal gold-based immunochromatographic assay for detection of ricin. Toxicon. 2002, 40: 255.
[110] Darby SM, Miller ML, Allen RO. Forensic determination of ricin and the alkaloid marker ricinine from castor bean extracts. J Forensic Sci. 2001 ,46{5):1033-42
[111] Despeyroux D, Walker N, Pearc M, et al. Characterization of ricin heterogeneity by electrospray mass spectrometry, capillary electrophoresis, and resonant mirror.Anal Biochem. 2000, 279(1):23-36
[112] Na DH, Cho CK, Youn YS, et al. Capillary electrophoresis to characterize ricin and its subunits with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Toxicon. 2004, 43(3): 329-335
[113] Rodriguez-Saona LE, Fry FS, Calvey EM. Use of fourier transform near-infrared reflectance spectroscopy for rapid quantification of castor bean meal in a selection of flour-based products. J Agric Food Chem. 2000, 48(11):5169-77
[114] Darby S. M, Miller M. L, Allen R. O. Forensic determination of ricin and the alkaloid marker ricinine from castor bean extracts. J.Forensic Sci. 2001, 46(5): 1033-1042.
[115] Narang, U,Anderson, G. P, Ligler, F. S, et al. Fiber optic-based biosensor for ricin. Biosens.Bioelectron. 1997, 12: 937-945.
[116] Ligler, F. S, Taitt, C. R, Shriver-Lake, L. C, et al. Array biosensor for detection of toxins. Anal.BioanaL Chem. 2003, 377: 469-477.
[117] Kirby R, Cho E. J, Gehrke B, et al. Aptamer-Based sensor arrays for the detection and quantitation of proteins. Anal. Chem. 2004, 76: 4066-4075.
[118] Taitt C. R, Anderson G. P, Lingerfelt B. M. et al. Nine-analyte detection using an array-based biosensor. Anal. Chem. 2002, 74: 6114-6120.
[119] 闫妍.蓖麻毒素和相思子毒素的提取纯化及基于生物质谱技术鉴定方法的建立. 硕士学位论文 2005
[120] 唐吉军.蓖麻毒素寡核苷酸适配子筛选方法的比较研究 硕士学位论文 2005
[121] 王顺涛. 重组蓖麻毒素A拮抗分子的设计 硕士学位论文 2005
[122] Godal A, Olsnes S, Pihl A. Ridioimmunoassay of abrin and ricin in blood. J Toxicol Environ Health. 1981,8(3): 409-17
[123] Shyu RH, SHyu HF, liu HW, et al. Colloidal gold-based immunochromatographic assay for detection of ricin. Toxicon 2002, 40(11): 255-8
[124] Griffiths GD, Newman H, Gee DJ. Identification and quantification of ricin toxin in animal tissues using ELISA. J Forensic Sci Soc. 1986, 26(5): 349-58.
[125] Leith Aq Griffiths GD, Green MA. Quantification of ricin toxin using a highly sensitive avidin/biotin enzyme-linked immunosorbent assay. J Forensic Sci Soc.1988, 28(4):227-36.
[126] Ramakrishnan S, Eagle MR, Houston LL. Radioimmunoassay of ricin A-and B-chains applied to samples of ricin A-chain prepared by chromatofocusing and by DEAE Bio-Gel A chromatography. Biochem Bivphys Acta. 1982, 719:341
[127] Sano T,Smith CL.Immuno-PCR:very sensitive antigen detection by means of specific antibody-DNA conjugates[J].Sciense,1992,258(5097): 120-122.
[128] 宋云扬,刘娟等.夹心免疫PCR方法检测蓖麻毒素[J]. 免疫学杂志 2002,18(3):229-231
[129] 严华.疫层析技术的应用与展望[J]. 微生物学免疫学进展.2005,33(3): 86-89
[130] Rong-Hwa Shyu,Huey-Fen Shyu.Colloida gold-based immunochromatographic assay for detection of ricin.[J]. Toxicon 2002,40: 255-258
[131] 汪琳,周琦等.免疫胶体金技术及其在检验检疫中的应用[J]. 检验检疫科学,2004,14(4):56-58
[132] 钱小红,谢剑炜.现代仪器分析在生物医学研究中的应用[J]. 化学工业出版社.2003,第一版,236
[133] 闫妍, 张淑珍等.一种基于肽质量指纹谱技术鉴定蓖麻毒素的新方法[J]. 分析化学研究报告,2006,34(2): 187-190
[134] 余涛,谢剑炜等.毒素的光纤免疫传感器研究进展[J]. 分析化学研究报告. 2004,32(6): 821-826
[135] U Narang, G P Anderson, F S Ligler et al.Biosens Bioelectron.1997,12(9-10): 937～945.
[136] G P Anderson ,KD King ,KL Gaffney et al.Biosens Bioelectron, 2000, 14 (10-11): 771-777.
[137] C A Rowe2Taitt, J W Hazzard, K E Hoffman et al. Biosens Bioelectron, 2000, 15(11-12): 579～589.
[138] 闫妍, 顾明松等.生物毒素的分析[J]. 化学通报, 2005,(4):285-290
[139] 朱立平等.免疫学常用实验方法.人民军医出版社.2000, 3
[140] 杨丽华,吕玉梅.SPR-生物传感器及其在食品安全与兽药残留检测中的研究进展.畜牧与饲料科学. 2007.5:38-40
[141] Rich R L ,Myszka D G. Advances in surface plasmon resonance biosensor analysis. Current Opinion in Biotechnology, 2000, 11(1):54
[142] Rich R L ,Myszka D G. Survey of the year 2000 commercial optical iosensor literature. Journal of Molecular ecognition ,2001, 14: 273
[143] 程惫,黄朝峰,段子渊.SPR生物传感器及其应用进展.中国生物工程杂志[J]. 2003, 5
[144] Naimushin, Alexei N. Soelberg. A portable surface plasmon resonance (SPR) sensor system with temperature regulation. Sensors and Actuators B:Chemical. 2003, 96(1-2):253-260
[145] Louis Amor Obando. Design manufactrure and application of fiber-optic surface plasmon resonance dip-probes. Arizona state university, 2001
[146] 陈军.光学电磁理论.科学出版社,2005
[147] 张雪娇.基于SPR传感技术的生物检测方法与试验研究.硕士学位论文.2007
[148] CHEN Li-hua,SU Zhong-min,WANG Rong-shun Summary of the investigation of the interaction between bio-molecules using surface plasmon resonance Journa of Molecular Science 2004,12: 44-49
[149] Rich R L ,Myszka D G. BIACORE J : a new platform for routine iomolecular interaction analysis. Journal of Molecular Recognition ,2001 ,14: 223-228
[150] Sunil K. Arya, Pratima R. Solanlti. S.P Singh. Poly-(3-hexylthiophene) self-assembled monolayer based cholesterol biosensor using surface plasmon resonance technique. Biosensors and Bioelectronics, 2007,22(11): 2516-2524
[151] Limin Cao, Hong Lin and Vladimir M. Mirsky. Surface plasmon resonance biosensor for enrofloxacin based on deoxyribonucleic acid. Analytica ChimicaActa, 2007,589(1): 1-5
[152] John Waswa, Joseph Irudayaraj and Chitrita DebRoy. Direct detection of E. Coli 0157:H7 in selected food systems by a surface plasmon resonance biosensor .LWT-Food Science and Technology,2007,40(2): 187-192
[153] Nylander C,L iedberg B, L ind T. Gas detection by means of surface p lasmon resonance. Senso rs &A ctuato rs, 1982, 3: 79-88.
[154] Stefan Lofaas, Malmqvist M , Ronnberg I, Stenberg E et al. Bioanalysis with surface palsmon resonance. Sensors & Actrators,1991, 5: 79-84.
[155] Kretschmann, E., The determination of the optical constants of metals by the excitation of surface plasmons. Z. Physik, 1971.241: 313-324.
[156] Otto, A., Excitation of surface plasma waves in silver by the method of frustrated total reflection. Z. Physik, 1968. 216: 398-410.
[157] Abeles, F. Surf. Sci,1976. 56: 237-251.
[158] Abeles, F. T. Lopez-Rios, A. Tadieddine, Solid State Commun, 1976,16: 843-847.
[159] Liedberg, B. C. Nylander, I. Lundstrom, Surface plasmons resonance for gas detection and biosensoring. Sensors and Actuators, 1983. 4: 299-304.
[160] 吴蕾.SPR生物传感器用于DNA聚合酶与DNA结合的研究.博士学位论文,2005
[161] 段媛媛,杨成丽.基于SPR生物传感器的免疫学检测.生物技术通讯2002,13,4: 264-268
[162] Indvk H E, Persson B S, Caselungbc M C, et al. Detemination of vitamin B12 in milk products and sclected foods by optical biosensor protein- binding assay: method comparison[J]. AOAC Int,2002,85(1): 72- 81
[163] Grace T, Stenberg E. Using surface plasmon resonance to determine vitamin concentrations [J]. Cereal Foods World,2002,(47): 7-9
[164] T A Grace, M Bostrom, B Persson, et al. Biosensor- Optical based assays- The determination of water- soluble vitamins in a verity of matrices by Biacore Q assay kits [J].Chem Bio Chem,2005,7(6):1196-1203
[165] 缪璐,张水华,刘仲明. SPR传感器在食品检测中的应用[J]. 食品科技2006,8: 266-279
[166] NAM IRA E J , HAYASH IDA T. Surface p lasmon resonance sensor for detection of some biomolecules [ J ].Sensors and Actuators, 1995:142-144.
[167] LAM W W, CHU L H. A surface p lasmon resonance system for the measurement of glucose in aqueous solution[J]. Sensors and Actuators 2003, 90: 264-270.
[168] 苏二辉,耿国强. 表面等离子共振生物传感技术在食品安全快速检测中的应用研究进展[J]. 河南农业大学学报,2007, 4: 113-116
[169] Rasooly A. Surface plasmon analysis of staphylococcal enterotoxin B in food[J]. Food Prot,2001,64(1):37-43
[170] Naimushin A N, Soelberg S D,Nguyen D K,et al. Detection of Staphyloccus aureus enterotoxin B at femtomolar levers with a miniature integrated two- channel surface plasmon resonance(SPR) sensor[J]. Biosens Bioelectron,2002,17(6-7):573-584
[171] 向四海,崔大付,蔡浩原等.利用表面等离子体谐振技术检测金黄色葡萄球菌肠毒素B[J]. 中国实验诊断学,2003,7(3):190-194
[172] Van der Gaag B, Spath S, Dietrich H,et al. Biosensors and multiple mycotoxin analysis[J]. Food Control,2003,(14): 251-254
[173] Gertie C. Immunochemical detection of Salmonella group B,D and E using an optical surface plasmon resonance biosensor [J]. FEMS Microbiology Letters,2003,(222): 197-200
[174] 葛晶,殷涌光,王凯.使用SPR生物传感器快速检测大肠杆菌E Coli 0157:H7[J]. 吉林大学学报(工学版), 2005,35(2): 214-218
[175] Mifumi Shimomur, Yoko Nomur, Wei Zhang,et al.Simple and rapid detection method using surface plasmon resonance for dioxins, polychlorinated biphenylx and atrazine[J]. Analytica Chimica Acta,2001,(434):223-230
[176] Ferguson JP, Baxter GA, McEvoy JDG, et al. Detection of streptomycin and dihydrostreptomycin residues in milk,honey and meat samples using an optical biosensor[J]. Analyst,2002,(127): 951-956
[177] Cacciatore G, Petz M, Rachid S, et al.Development of an optical biosensor assay for detection of lactam antibiotics in milk using the penicillin- binding proteinx[J]. Analytica Chimica Acta,2004,(520): 105-115
[178] Tschmelak J, Proll G,GauglitzG. Optical biosensor for pharmaceuticals,antibiotics, hormones, endocrine disrupting chemicals and pesticides in water:Assay optimization process for estrone as example[J]. Talanta,2005,(65): 313-323
[179] Samsonova J Baxter GCrooks S, et al. Biosensor immunoassay of ivermectin in bovine milk[J]. AOAC Int,2002,(85):879-882
[180] Crooks SRH, McCarney B, Traynor IM, et al. Detection of levamisole residues in bovine liver and milk by immunobiosensor[J]. Anal Chim Acta,2003,(483):181-186
[181] Haasnoot W, Bienenmann - Ploum M, Kohen F. Biosen-sorimmunoassay for the detection of eight sulfonamides in chicken serum[J]. Anal Chim Acta,2003,(483):171-180
[182] Shelver W, Smith DJ. Determination of ractopamine in cattle and sheep urine samples using an optical biosensor analysis: comparative study with HPLC and ELISA[J]. Agric Food Chem,2003,(51): 3715-3721
[183] CHAH S, YI J , ZARE R N. Surface p lasmon resonance analysis of aqueous mercuric ions [ J ]. Sensors and Actuators, 2004. 99: 216-222.
[184] JORN C C, YU A, EDWARD P C, et al. Surface plasmon resonance sensor for Hg ( Ⅱ) detection by binding interactions with polypyrrole and 22mercap tobenzothiazole[ J ]. Sensors and Actuators, 2004. 101: 236 - 241.
[185] D IASF ILHO N L, GUSH IKEM Y, POL IlTO W L. 22Mercap tobenzothiazole clay as matrix for sorp tion and preconcentration of some heavy metals from aqueous solution[ J ]. Anal Chim Acta, 1995, 306: 167-172.
[186] KIM S H, HAN S K, KIM J H, et al. A self2assembled squarylium dye monolayer for the detection ofmetal ions by surface plasmon resonances [ J ]. Dyes Pigments,2000, 44: 55-61.
[187] NELSONRW, KRONEJR. Advances in surface plasmon resonance bimolecular interaction analysis mass spectrometry (BIA/MS) [J]. Mol Recognit ,1999 ,12 (2): 77-93
[188] ROBBIO L L ,UBOLDI P ,MARCOVINA S, et al. Epitope mapping analysis of apolipoprotein B-100 using a surface plasmon resonance-based biosensor [J]. Biosens Bioelectron ,2001,16 (9-12): 963-969.
[189] WERAWATGOOMPAS ,SRIYUDTHSAKM. Feasibility study on the detection of ferritin using surface plasmon resonance [J]. J Med Assoc Thai ,2001 ,84 (Suppl 1):155-162.
[190] 孙颖,张阳德.表面等离子共振技术在蛋白质组学中的应用[J]. 医学与哲学,2002,23:30-32