近红外荧光试剂Cy系列的合成研究
|
摘要
荧光标记技术起源于20世纪40年代,随着现代医学、分子生物学的发展和各种先进荧光检测技术及仪器的应用,荧光标记作为一种非放射性的标记技术受到很大程度的重视,并取得了迅速的发展,广泛应用于细胞内外物质检测、核酸的检测及疾病早期诊断等。近红外荧光试剂Cy系列是不同杂环核通过甲川链连接起来的一类不饱和杂环化合物,在检测及定量标记组分时具有很高的消光系数及量子产率,成为基因芯片测试用的首选荧光标记物。Cy系列试剂虽然已商品化,如Cy3、Cy5、Cy5.5,但价格昂贵,使得其应用受到了限制。鉴于Cy系列试剂的广泛用途和应用前景,探索低成本,高收率的合成方法,无疑会拓宽Cy系列荧光试剂在国内的应用,促进生物学和医药学的发展。本文探讨了生物荧光分析试剂Cy系列的合成研究,报告了该研究的思路和取得的成果。
本文第一章首先综述了常见的荧光标记试剂以及它们的优缺点,重点介绍了Cy系列试剂的结构特点、光谱特征、光稳定性及水溶性。由于Cy系列试剂的特殊结构使其在荧光标记方面与其他试剂相比有着显著的优点,在生物芯片和蛋白质组学方面得到了广泛的应用,因此文章综述了Cy系列试剂在DNA芯片和蛋白质组学研究中的标记方式以及应用。
第二章探讨了对肼基苯磺酸的合成工艺。对肼基苯磺酸是由对氨基苯磺酸经亚硝酸钠重氮化得到的重氮盐用亚硫酸氢钠还原、水解、硫酸酸化制得,本文经多次实验确定了反应原料、用量及反应条件,得到的产品纯度好,产率高。
论文第三章合成了β-苯胺基丙烯醛缩苯胺。该研究以丙炔醇为原料,经三氧化铬氧化生成丙炔醛后,直接与苯胺反应简便地合成了β-苯胺基丙烯醛缩苯胺,并通过正交实验的方法优化了反应条件。得出最佳反应条件为:6.3mL(0.10mol)丙炔醇中加入8.5g(0.085mol)三氧化铬的硫酸水溶液,氧化反应2小时后,有机相在冰浴冷却下再与25mL(0.25mol)苯胺和63mL10%盐酸反应得到β-苯胺基丙烯醛缩苯胺盐酸盐,收率为48.4%。将该盐酸盐溶于甲醇与氢氧化钠溶液作用得β-苯胺基丙烯醛缩苯胺。
论文第四章合成N,N′-二苯基甲脒,本合成法以甲酸与苯胺为原料,硼砂为催化剂,并在脱水剂的存在下,经两步脱水反应而得二苯甲脒盐酸盐,再经碱化得最终产品。本文还通过~1H NMR跟踪反应,确定了合成二苯甲脒的最佳反应时间,在不影响反应收率的前提下,大大缩短了反应时间。
With the developments of medicine, molecular biochemistry and various advanced fluorescence detecting techniques, fluorescence labeling technology, a non-radioactive labeling technology, has been becoming more and more attractive due to its broad and potential applications in protein and nucleic acid assay, poison determination and clinical medical diagnose. Cy series reagents are the first selected fluorescence probe in biological analysis because they have many advantages, such as the better extinct coefficient and the higher quantum efficiency. However, the usages of Cy series has been limited in our country because of their high price. It is crucial to design and seek for a low costs but high yield synthetic procedure of Cy series. So our group studies this problem. The researches and results are reported in the paper.In this paper, the general fluorescence reagents are been first reviewed. Especially, it focuses on presenting the structure, spectroscopic character, light stability and dissolubility of Cy series. According to the properties of Cy series, it is be widely used in proteomics and DNA chip. The methods to label Cy reagents to protein and nucleic acid are mentioned and the applications in proteomic and DNA chip are illustrated.In the second chapter, p-hydrazino-benzenesulfonic acid was prepared by diazotizing sulfanili acid with sodium nitrite. The diazanium salt was then reduced with sodium hydrogen sulfite and acidified with sulfuric acid to give 83.4% product. After several repeated experiments, the reactant and reaction condition are determined. The purity of the compound is very good and the yield is quite high.In the third part of this paper, a facile method is presented for synthesis of malonaldehyde dianil from propargyl alcohol by oxidizing using chromium trioxide and addition reaction with aniline. In this research, the best reaction condition was obtained by orthogonal tests. 6.3mL(0.10mol)propargyl alcohol was oxidized with 8.5g (0.085mol)chromium trioxide. After oxidizing for two hours, the organic layer was separated and diluted with 200mL water. 25mL(0.25mol)aniline and 63mL 10% hydrochloric acid were added, respectively, under the ice-bath condition. The yield of malonaldehyde dianil hydrochloride is 48.4%. Malonaldehyde dianli was obtained by
本文第一章首先综述了常见的荧光标记试剂以及它们的优缺点,重点介绍了Cy系列试剂的结构特点、光谱特征、光稳定性及水溶性。由于Cy系列试剂的特殊结构使其在荧光标记方面与其他试剂相比有着显著的优点,在生物芯片和蛋白质组学方面得到了广泛的应用,因此文章综述了Cy系列试剂在DNA芯片和蛋白质组学研究中的标记方式以及应用。
第二章探讨了对肼基苯磺酸的合成工艺。对肼基苯磺酸是由对氨基苯磺酸经亚硝酸钠重氮化得到的重氮盐用亚硫酸氢钠还原、水解、硫酸酸化制得,本文经多次实验确定了反应原料、用量及反应条件,得到的产品纯度好,产率高。
论文第三章合成了β-苯胺基丙烯醛缩苯胺。该研究以丙炔醇为原料,经三氧化铬氧化生成丙炔醛后,直接与苯胺反应简便地合成了β-苯胺基丙烯醛缩苯胺,并通过正交实验的方法优化了反应条件。得出最佳反应条件为:6.3mL(0.10mol)丙炔醇中加入8.5g(0.085mol)三氧化铬的硫酸水溶液,氧化反应2小时后,有机相在冰浴冷却下再与25mL(0.25mol)苯胺和63mL10%盐酸反应得到β-苯胺基丙烯醛缩苯胺盐酸盐,收率为48.4%。将该盐酸盐溶于甲醇与氢氧化钠溶液作用得β-苯胺基丙烯醛缩苯胺。
论文第四章合成N,N′-二苯基甲脒,本合成法以甲酸与苯胺为原料,硼砂为催化剂,并在脱水剂的存在下,经两步脱水反应而得二苯甲脒盐酸盐,再经碱化得最终产品。本文还通过~1H NMR跟踪反应,确定了合成二苯甲脒的最佳反应时间,在不影响反应收率的前提下,大大缩短了反应时间。
With the developments of medicine, molecular biochemistry and various advanced fluorescence detecting techniques, fluorescence labeling technology, a non-radioactive labeling technology, has been becoming more and more attractive due to its broad and potential applications in protein and nucleic acid assay, poison determination and clinical medical diagnose. Cy series reagents are the first selected fluorescence probe in biological analysis because they have many advantages, such as the better extinct coefficient and the higher quantum efficiency. However, the usages of Cy series has been limited in our country because of their high price. It is crucial to design and seek for a low costs but high yield synthetic procedure of Cy series. So our group studies this problem. The researches and results are reported in the paper.In this paper, the general fluorescence reagents are been first reviewed. Especially, it focuses on presenting the structure, spectroscopic character, light stability and dissolubility of Cy series. According to the properties of Cy series, it is be widely used in proteomics and DNA chip. The methods to label Cy reagents to protein and nucleic acid are mentioned and the applications in proteomic and DNA chip are illustrated.In the second chapter, p-hydrazino-benzenesulfonic acid was prepared by diazotizing sulfanili acid with sodium nitrite. The diazanium salt was then reduced with sodium hydrogen sulfite and acidified with sulfuric acid to give 83.4% product. After several repeated experiments, the reactant and reaction condition are determined. The purity of the compound is very good and the yield is quite high.In the third part of this paper, a facile method is presented for synthesis of malonaldehyde dianil from propargyl alcohol by oxidizing using chromium trioxide and addition reaction with aniline. In this research, the best reaction condition was obtained by orthogonal tests. 6.3mL(0.10mol)propargyl alcohol was oxidized with 8.5g (0.085mol)chromium trioxide. After oxidizing for two hours, the organic layer was separated and diluted with 200mL water. 25mL(0.25mol)aniline and 63mL 10% hydrochloric acid were added, respectively, under the ice-bath condition. The yield of malonaldehyde dianil hydrochloride is 48.4%. Malonaldehyde dianli was obtained by
引文
[1] 李宁,许红韬,蛋白质组研究的现状与展望,生物技术通讯,2000,11(14),281-285.
[2] 严凤霞,王莜敏,现代化学仪器分析选论,上海:华东师范大学出版社,1992.221.
[3] A. Rudolf, L. Wolfgang, Ribosomal RNA-targeted nucleic acid probes for studies in microbial ecology, FEMS Micorobiol. Rev. , 2000, 24, 555-565.
[4] M. A. King, Detection of dead cells and measurement of cell killing by flow cytometry, J. Immunol. Methods, 2000, 243, 155-166.
[5] H. M. Shapiro, Microbial analysis at the single-cell level: tasks and techniques, J. Microbiol. Methods, 2000, 42, 3-16.
[6] 匡春香,权春善,范圣第,含活性基荧光染料及其在生物技术中的应用,染料工业,1997,34(2),16-20.
[7] 吕荣文,张淑芬,杨锦宗,化学分析用荧光染料,染料工业,1999,36(6):14-17.
[8] 朱爱平,吴世康,香豆素化合物在不同溶剂中荧光研究,物理化学学报:1998.14(6),552-556.
[9] R. P. Haugland, Handbook of fluorescent probes and research products, 9th edition, Molecular Probes, Eugene, OR, USA, 2002.
[10] K. Yamana, Y. Ohashi, K. Nunota, H. Nakano, Synthesis, binding and fluorescence properties of oligonucleotide derivatives having a dansyl fluorescence label attached to the 2'-position of a ribonucleoside, Tetrahedron, 1997, 53, 4265-4270.
[11] 叶云,浅谈菁染料及其研究前景,大学化学,1999,14(3),55-58.
[12] 郁美娟,孟庆华,任吉存,基于生命体系中氨基键合标记的荧光探针研究进展,染料与染色,2004,41(3),137-144.
[13] D. C. Prasher, Gene 111, 1992, 229-233.
[14] 汪恒英,周守标,常志州,绿色荧光蛋白(GFP)研究进展,生物技术,2004,14(3),71-73.
[15] 杨祥宇,宋健,冯荣秀,荧光标记染料,化学通报,2003,9,615-621.
[16] 王丽秋,彭孝军,生物标示用3H-吲哚菁染料,染料工业,2002,39(4),8-12.
[17] 曾万学,陈萍,盛丽琴,单重态氧淬灭剂对长链菁染料的光稳定作用,中国 科学,1995,25(6),579-584.
[18] 赵江,陈萍,李军,菁染料薄膜的光谱性能及稳定性的研究,感光科学与光化学,1997,15(1),59-65.
[19] 孙树清,陈萍,可录型光盘与菁染料光存储媒体的研究进展,化学通报,1999,6,20-26.
[20] 李军,陈萍,赵江,近红外吸收菁染料分子链结构对其光氧化稳定性能的影响,感光科学与光化学,1997,15(4),343-352.
[21] 俞开潮,金玲,程红,七甲川花菁染料的研究进展,合成化学,2004,12,38-42.
[22] 郑洪,李东辉,一种近红外花菁染料的合成及其应用于生物大分子的测定,应用化学,1999,16(3),7-20.
[23] Y. Inaguma, J. H. Sohn, S. Kiml, Quantum paraelectricityina perovskite La_(0.5) Na_(0.5) TiO_3, J. Phys. Soc. Jphn. , 1992, 61, 3831-3832.
[24] A. S. Waggoner, Cyanine dyes as labeling icaland other materials by luminescence method, US Patent, 5627027, 1997, 05, 06.
[25] R. B. Mujumdar, L. A. Earnl, S. R. Mujumdar, C. J. Lewis and A. S. Waggoner, Cyanine dye labeling reagents: sulfoindocyanine succinimidyl esters, Bioconjugate Chem. , 1993, 4, 105-111.
[26] 车庆林,彭孝军,荧光免疫分析用染料及其研究进展,现代化工,1998,2,12-14.
[27] A. T. Woltey, G. F. Sensabaugh, High-speed DNA genotyping using microfabricated capillary array electrophoresis chips, Anal. Chem. , 1997, 69 (11), 2181-2186.
[28] S. M. Clak, R. A. Mathies, Multiplex dsDNA fragment sizing using dimeric intercalation dyes and capillary array electrophoresis: ionic effects on the stability and electrophoretic mobility of DNA-Dye complexes, Anal. Chem. , 1997, 69 (7), 1355-1363.
[29] H. Su-Chun, J. Jingyue, Cyanine dyes with high absorption cross section as donor chromophores in energy transfer primers~1, Anal. Biochem. , 1996, 243, 15-27.
[30] K. Narasimhachari, P. Gabor, A new method for the synthesis of heptamethine cyanine dyes: synthesis of new near-infrared fluorescent labels. , J. Org. Chem. , 1995, 60 (8), 2391-2395.
[31] 陈中举,张燕玲,黄金瑛,荧光标记生物大分子及其应用,国外医学生物医学工程分册,2004,27(6),348-352.
[32] B. Andrea, G. Andera, P. Giovarma, Use of probes for detecting DNA single base mutation by capillary electrophoresis, Electrophoresis, 2002, 23 (6), 926-929.
[33] H. Haiyong, D. J. Bearss, L. W. Browne, Ideneification of differentially expressed gene sinpan creatic cancer cells using cDNA microarray, Cancer Res. , 2002, 62 (1), 2890-2896.
[34] 丁·格里菲思著,侯毓汾,吴祖望译,颜色与有机分子结构,北京:化学工业出版社,1985,254-284.
[35] 皮格拉夫基德,照相化学(第四分册:光学敏化),刘敦译,北京:中国电影出版社,1984,1,4.
[36] U. Schobel, H. J. Egelhaaf, A. Brechl, D. Oelkmg, G. Gauglitz, A new donor-acceptor pair for fluorescent immunoassays by energy transfer, Bioconjugate Chem. , 1999, 10 (6), 1107-1114.
[37] S. R. Mujumbdar, R. B. Mujumbdar, C. M. Grant and A. S. Waggoner, Cyanine-labeling reagents: sulfobenzindocyanine succinimidyl esters, Bioconjugate Chem. , 1996, 7 (3), 356-362.
[38] G. Ramsay, DNA Chips: State of the art, Nat. Biotechnol. , 1998, 16 (1), 40-44.
[39] Q. Y. Wu, Foundation Life Science, Beijing: Higher Education Press, 2002, 248-249.
[40] Z. Guo, R. A. GuilfOyle, A. J. Thiel, Direct fluorescence analysis of genetic polymorphisms by hybridization with oligonucleotide arrays on glass supports, Nucleic Acids Res. , 1994, 22, 5456-5465.
[41] 王璐,基因芯片技术研究现状,医学动物防制,2005,21(1),3-6.
[42] 刘和平,王宏,陆祖宏,一种基于寡核苷酸微阵列芯片的多重可扩增探针杂交技术,遗传学报,2004,31(2),119-124.
[43] 张卫,王米渠,施茵,应用基因芯片研究针刺对亚急性衰老大鼠蛋白质合成相关基因表达的影响,江西中医学院学报,2004,16(4),53-56.
[44] K. Ruscher, A fluorescence based non-radioactive lectrophoretic mobility shift assay, J. Biotechnol. , 2000, 78, 163-170.
[45] 李成涛,寡核苷酸DNA芯片用于ABO血型基因分型的研究,临床检验杂志,2002,20(3),139-141.
[46] 冒海蕾,崔之础,施健,基因芯片在差异表达研究中测定影响因素的初步探讨,南通医学院学报,2004,24(2),159-161.
[47] 祝骥,马文丽,李凌,基因表达谱芯片杂交影响因素的初步研究,生命科学研究,2004,8(2),133-139.
[48] S. Bupinder, A. J. Bhar, Rapid and accurate determination of (CAG)n repeats in the androgen receptor gene using polymerase chain reaction and automated fragment analysis, Clin. Biochem. , 1999, 32 (5), 327-332.
[49] H. Z. Xing, Low-level arsenite induced, gene expression in HEK293 cells, Toxicology, 2003, 187, 39-48.
[50] 宋家武,检测拉米夫定耐药位点基因芯片的研制及其应用初探,中华检验医学杂志,2003,26(2),83-85.
[51] 徐宜英,任莉莉,基因芯片技术检测拉美夫定相关的HBV变异,中国病毒学,2004,19(4),410-412.
[52] 陈忠斌,应用抑制消减杂交和cDNA芯片技术筛选流行性感冒病毒感染相关基因,病毒学报,2003,19(1),21-26.
[53] S. Frank, Automated image analysis and in situ hybridization as tools to study bacterial populations in food resources, gut and cast of lumbricus terrestris L, J. Microbiol. Methods, 2002, 48, 53-68.
[54] 纪建国,茹炳根,蛋白质组学研究相关技术及其在生物医学研究中的应用,药物生物技术,2002,9(1),1-11.
[55] 万晶宏,贺福初,蛋白质组技术的研究进展,科学通报,1999,44(9),904-911.
[56] 薛淑群,刘伟,匡友谊,蛋白质组学研究的技术体系及进展,水产学杂志,2004,17(1),77-81.
[57] 李兆英,蛋白质组学及其意义,生物学通报,2001,35(9),8-9.
[58] 洪龙,廖玮,魏芳,用于识别不同细胞蛋白质组的噬菌体抗体芯片,物理化学学报,2004,20(10),1182-1185.
[59] G. Macbeath, Printing proteins as microarrays for high-throughput function determination, Science, 2000, 289, 1760.
[60] 郑洪,李东辉,陈秋影,许金钩,一种近红外花菁染料的合成及其应用于生物大分子的测定,应用化学,1999,3,17-20.
[61] R. J. Williams, J. M. Peralta, V. W. Tazng, Near infrared heptamethine cyanine dyes: a new tracer for solid phase immunoassays, Appl. Sepectrosc., 1997, 51, 1435-1435.
[62] J. Sowwell, A. K. Heard, J. C. Mason, Use of noncovalent labeling in illustracting ligand binding to human serum album in via affinity capillary electroporesis with near infred laser induced fluorescence detection, J. Chromatogra. B, 2001, 755, 91-99.
[63] R. J. Bram, In vivo monitoring of the distribution of a monoclonal antibody fragment, Vib. Spectrosc. , 2002, 28, 177-188.
[64] C. M. Liu, Analysis of secretory immunoglobulin in human saliva by laser-induced fluorescence capillary electrophoresis, J. Chromatogra. B, 2003, 791, 315-321.
[65] S. Kamlakshi, Post-column reaction detection based on fluorescence energy transfer in the far red spectral region, J. Pharm. Biomed. Anal. , 1999, 21, 361-370.
[66] 谢文章,蛋白质微阵列检测抗原—抗体相互作用,生物化学与生物物理进展,2002,29(2),311-315.
[67] 张庆峰,高志贤,王升启,用于雌二醇检测的免疫芯片技术,中国生物工程杂志,2004,24(9),85-88.
[68] 马增春,高月,吕俊萍,双向凝胶电泳中三种蛋白质检测方法的比较,中国生物化学与分子生物学报,2004,20(4),551-556.
[69] R. Redkar, Real-time detection of brucella abortus, brucella melitensis and brucella suis, Mole. Cell. Probes, 2001, 15, 43-52.
[70] Y. Ishii, Fluorescence resonance energy transfer between single fluorophores attached to a coiled-coil protein in aqueous solution, Chem. Phys. , 247, 1999, 163-173.
[71] R. Yamasakil, Single molecular observation of the interaction of GroEL with substrate proteins, J. Mol. Biol. , 1999, 292, 965-972.
[72] Y. K. Suzukia, Imaging of the fuorescence spectrum of a single fluorescent molecule by prism-based spectroscopy, FEBS Letters, 2002, 512, 235-239.
[73] 杨军,N-十二烷基-N′-(对氨基苯磺酸钠)硫脲的合成,现代化工,1999,19(6),26-27.
[74] 徐克勋,精细有机化工原料及中间体手册,北京:化学工业出版社,3-621.
[75] 马世昌,基础有机化学反应,西安:陕西人民教育出版社,1995,第一版,665.
[76] R. H. Munchen, uber malodialdehyde I, mitteilung, Ber. , 1941, 74 (3), 1825-1829.
[77] S. R Williamt, Two simple amidinium vinylogs, J. Am. Chem. Soc. , 1949, 71, 754-755.
[78] G. L. Stagno, D. Alcontres, G. Lovecchio, Influence of conjugation on the oxidation of alcohols with selenium dioxide, Gazz. Chim. Ital. , 1960, 90, 337-346.
[79] 樊能廷,有机合成事典,北京:北京理工大学出版社,1992,665-666.
[80] M. G. Veliev, M. M. Guseinov, An imprc, ved synthesis of propynal, Synthesis, 1980, 6, 461.
[81] NIST Scientific and Technical Database, http://www.nist.gov/srd/nist35.ntm.
[82] 上海市科学技术交流站组编,正交试验设计法,上海:上海人民出版社,1975.18-23.
[83] B. Tom, H. R. Manchester and H. R. Ernest, United states Patent Office, 1933, 1, 933, 206.
[84] W. H. Warren and F. E. Wilson, Aaction of thionyl chloride on some anilides, ureas and urethane, Ber. 68B, 1935, 957-960.
[85] E. H. John, R. B. Alfred, Unsaturated aromatic amines; a novel synthesis of indoles, J. Am. Chem. Soc. , 1958, 80 (2), 437-439.
[86] P. Karel and G. Hans, A-Halo others, Ⅶ. reactions of dichloromethyl alkyl ethers with amino acid derivatives, Chem. Ber. , 1961, 49, 527-537.
[87] R. Debrath, A. Worth, Symmetrically disubstituted formamidines, formation by raney-nicked rudiction of thioureas, J. Chem. Soc. , 1948, 1716-1720.
[88] M. Passerini, The isonitriles Ⅳ. reaction with organic acids, Gazz. Ber. Ital. , 1922, 52 (Ⅱ), 250-257.
[89] R. M. Roberts, Ortho ester, imilic ester and amidimes, the identity of N-phenyl-N'-p-toylformamidine, J. Am. Chem. Soc. , 72, 3603-3610.
[90] 王晓峰,发明专利申请公开说明书,申请号:89105167.8.公开号CN:1050377A.
[91] 马立人,蒋中华,生物芯片,北京:化学工业出版社,2000,3,11.
[92] 楼开炎,不对称水溶性菁染料的简便制备与纯化方法,华东理工大学学报,2002,28(2),212-215.
[93] W. Harold, A synthesis of bromocaproic acid, J. Am. Chem. Soc. , 1951, 73,
[2] 严凤霞,王莜敏,现代化学仪器分析选论,上海:华东师范大学出版社,1992.221.
[3] A. Rudolf, L. Wolfgang, Ribosomal RNA-targeted nucleic acid probes for studies in microbial ecology, FEMS Micorobiol. Rev. , 2000, 24, 555-565.
[4] M. A. King, Detection of dead cells and measurement of cell killing by flow cytometry, J. Immunol. Methods, 2000, 243, 155-166.
[5] H. M. Shapiro, Microbial analysis at the single-cell level: tasks and techniques, J. Microbiol. Methods, 2000, 42, 3-16.
[6] 匡春香,权春善,范圣第,含活性基荧光染料及其在生物技术中的应用,染料工业,1997,34(2),16-20.
[7] 吕荣文,张淑芬,杨锦宗,化学分析用荧光染料,染料工业,1999,36(6):14-17.
[8] 朱爱平,吴世康,香豆素化合物在不同溶剂中荧光研究,物理化学学报:1998.14(6),552-556.
[9] R. P. Haugland, Handbook of fluorescent probes and research products, 9th edition, Molecular Probes, Eugene, OR, USA, 2002.
[10] K. Yamana, Y. Ohashi, K. Nunota, H. Nakano, Synthesis, binding and fluorescence properties of oligonucleotide derivatives having a dansyl fluorescence label attached to the 2'-position of a ribonucleoside, Tetrahedron, 1997, 53, 4265-4270.
[11] 叶云,浅谈菁染料及其研究前景,大学化学,1999,14(3),55-58.
[12] 郁美娟,孟庆华,任吉存,基于生命体系中氨基键合标记的荧光探针研究进展,染料与染色,2004,41(3),137-144.
[13] D. C. Prasher, Gene 111, 1992, 229-233.
[14] 汪恒英,周守标,常志州,绿色荧光蛋白(GFP)研究进展,生物技术,2004,14(3),71-73.
[15] 杨祥宇,宋健,冯荣秀,荧光标记染料,化学通报,2003,9,615-621.
[16] 王丽秋,彭孝军,生物标示用3H-吲哚菁染料,染料工业,2002,39(4),8-12.
[17] 曾万学,陈萍,盛丽琴,单重态氧淬灭剂对长链菁染料的光稳定作用,中国 科学,1995,25(6),579-584.
[18] 赵江,陈萍,李军,菁染料薄膜的光谱性能及稳定性的研究,感光科学与光化学,1997,15(1),59-65.
[19] 孙树清,陈萍,可录型光盘与菁染料光存储媒体的研究进展,化学通报,1999,6,20-26.
[20] 李军,陈萍,赵江,近红外吸收菁染料分子链结构对其光氧化稳定性能的影响,感光科学与光化学,1997,15(4),343-352.
[21] 俞开潮,金玲,程红,七甲川花菁染料的研究进展,合成化学,2004,12,38-42.
[22] 郑洪,李东辉,一种近红外花菁染料的合成及其应用于生物大分子的测定,应用化学,1999,16(3),7-20.
[23] Y. Inaguma, J. H. Sohn, S. Kiml, Quantum paraelectricityina perovskite La_(0.5) Na_(0.5) TiO_3, J. Phys. Soc. Jphn. , 1992, 61, 3831-3832.
[24] A. S. Waggoner, Cyanine dyes as labeling icaland other materials by luminescence method, US Patent, 5627027, 1997, 05, 06.
[25] R. B. Mujumdar, L. A. Earnl, S. R. Mujumdar, C. J. Lewis and A. S. Waggoner, Cyanine dye labeling reagents: sulfoindocyanine succinimidyl esters, Bioconjugate Chem. , 1993, 4, 105-111.
[26] 车庆林,彭孝军,荧光免疫分析用染料及其研究进展,现代化工,1998,2,12-14.
[27] A. T. Woltey, G. F. Sensabaugh, High-speed DNA genotyping using microfabricated capillary array electrophoresis chips, Anal. Chem. , 1997, 69 (11), 2181-2186.
[28] S. M. Clak, R. A. Mathies, Multiplex dsDNA fragment sizing using dimeric intercalation dyes and capillary array electrophoresis: ionic effects on the stability and electrophoretic mobility of DNA-Dye complexes, Anal. Chem. , 1997, 69 (7), 1355-1363.
[29] H. Su-Chun, J. Jingyue, Cyanine dyes with high absorption cross section as donor chromophores in energy transfer primers~1, Anal. Biochem. , 1996, 243, 15-27.
[30] K. Narasimhachari, P. Gabor, A new method for the synthesis of heptamethine cyanine dyes: synthesis of new near-infrared fluorescent labels. , J. Org. Chem. , 1995, 60 (8), 2391-2395.
[31] 陈中举,张燕玲,黄金瑛,荧光标记生物大分子及其应用,国外医学生物医学工程分册,2004,27(6),348-352.
[32] B. Andrea, G. Andera, P. Giovarma, Use of probes for detecting DNA single base mutation by capillary electrophoresis, Electrophoresis, 2002, 23 (6), 926-929.
[33] H. Haiyong, D. J. Bearss, L. W. Browne, Ideneification of differentially expressed gene sinpan creatic cancer cells using cDNA microarray, Cancer Res. , 2002, 62 (1), 2890-2896.
[34] 丁·格里菲思著,侯毓汾,吴祖望译,颜色与有机分子结构,北京:化学工业出版社,1985,254-284.
[35] 皮格拉夫基德,照相化学(第四分册:光学敏化),刘敦译,北京:中国电影出版社,1984,1,4.
[36] U. Schobel, H. J. Egelhaaf, A. Brechl, D. Oelkmg, G. Gauglitz, A new donor-acceptor pair for fluorescent immunoassays by energy transfer, Bioconjugate Chem. , 1999, 10 (6), 1107-1114.
[37] S. R. Mujumbdar, R. B. Mujumbdar, C. M. Grant and A. S. Waggoner, Cyanine-labeling reagents: sulfobenzindocyanine succinimidyl esters, Bioconjugate Chem. , 1996, 7 (3), 356-362.
[38] G. Ramsay, DNA Chips: State of the art, Nat. Biotechnol. , 1998, 16 (1), 40-44.
[39] Q. Y. Wu, Foundation Life Science, Beijing: Higher Education Press, 2002, 248-249.
[40] Z. Guo, R. A. GuilfOyle, A. J. Thiel, Direct fluorescence analysis of genetic polymorphisms by hybridization with oligonucleotide arrays on glass supports, Nucleic Acids Res. , 1994, 22, 5456-5465.
[41] 王璐,基因芯片技术研究现状,医学动物防制,2005,21(1),3-6.
[42] 刘和平,王宏,陆祖宏,一种基于寡核苷酸微阵列芯片的多重可扩增探针杂交技术,遗传学报,2004,31(2),119-124.
[43] 张卫,王米渠,施茵,应用基因芯片研究针刺对亚急性衰老大鼠蛋白质合成相关基因表达的影响,江西中医学院学报,2004,16(4),53-56.
[44] K. Ruscher, A fluorescence based non-radioactive lectrophoretic mobility shift assay, J. Biotechnol. , 2000, 78, 163-170.
[45] 李成涛,寡核苷酸DNA芯片用于ABO血型基因分型的研究,临床检验杂志,2002,20(3),139-141.
[46] 冒海蕾,崔之础,施健,基因芯片在差异表达研究中测定影响因素的初步探讨,南通医学院学报,2004,24(2),159-161.
[47] 祝骥,马文丽,李凌,基因表达谱芯片杂交影响因素的初步研究,生命科学研究,2004,8(2),133-139.
[48] S. Bupinder, A. J. Bhar, Rapid and accurate determination of (CAG)n repeats in the androgen receptor gene using polymerase chain reaction and automated fragment analysis, Clin. Biochem. , 1999, 32 (5), 327-332.
[49] H. Z. Xing, Low-level arsenite induced, gene expression in HEK293 cells, Toxicology, 2003, 187, 39-48.
[50] 宋家武,检测拉米夫定耐药位点基因芯片的研制及其应用初探,中华检验医学杂志,2003,26(2),83-85.
[51] 徐宜英,任莉莉,基因芯片技术检测拉美夫定相关的HBV变异,中国病毒学,2004,19(4),410-412.
[52] 陈忠斌,应用抑制消减杂交和cDNA芯片技术筛选流行性感冒病毒感染相关基因,病毒学报,2003,19(1),21-26.
[53] S. Frank, Automated image analysis and in situ hybridization as tools to study bacterial populations in food resources, gut and cast of lumbricus terrestris L, J. Microbiol. Methods, 2002, 48, 53-68.
[54] 纪建国,茹炳根,蛋白质组学研究相关技术及其在生物医学研究中的应用,药物生物技术,2002,9(1),1-11.
[55] 万晶宏,贺福初,蛋白质组技术的研究进展,科学通报,1999,44(9),904-911.
[56] 薛淑群,刘伟,匡友谊,蛋白质组学研究的技术体系及进展,水产学杂志,2004,17(1),77-81.
[57] 李兆英,蛋白质组学及其意义,生物学通报,2001,35(9),8-9.
[58] 洪龙,廖玮,魏芳,用于识别不同细胞蛋白质组的噬菌体抗体芯片,物理化学学报,2004,20(10),1182-1185.
[59] G. Macbeath, Printing proteins as microarrays for high-throughput function determination, Science, 2000, 289, 1760.
[60] 郑洪,李东辉,陈秋影,许金钩,一种近红外花菁染料的合成及其应用于生物大分子的测定,应用化学,1999,3,17-20.
[61] R. J. Williams, J. M. Peralta, V. W. Tazng, Near infrared heptamethine cyanine dyes: a new tracer for solid phase immunoassays, Appl. Sepectrosc., 1997, 51, 1435-1435.
[62] J. Sowwell, A. K. Heard, J. C. Mason, Use of noncovalent labeling in illustracting ligand binding to human serum album in via affinity capillary electroporesis with near infred laser induced fluorescence detection, J. Chromatogra. B, 2001, 755, 91-99.
[63] R. J. Bram, In vivo monitoring of the distribution of a monoclonal antibody fragment, Vib. Spectrosc. , 2002, 28, 177-188.
[64] C. M. Liu, Analysis of secretory immunoglobulin in human saliva by laser-induced fluorescence capillary electrophoresis, J. Chromatogra. B, 2003, 791, 315-321.
[65] S. Kamlakshi, Post-column reaction detection based on fluorescence energy transfer in the far red spectral region, J. Pharm. Biomed. Anal. , 1999, 21, 361-370.
[66] 谢文章,蛋白质微阵列检测抗原—抗体相互作用,生物化学与生物物理进展,2002,29(2),311-315.
[67] 张庆峰,高志贤,王升启,用于雌二醇检测的免疫芯片技术,中国生物工程杂志,2004,24(9),85-88.
[68] 马增春,高月,吕俊萍,双向凝胶电泳中三种蛋白质检测方法的比较,中国生物化学与分子生物学报,2004,20(4),551-556.
[69] R. Redkar, Real-time detection of brucella abortus, brucella melitensis and brucella suis, Mole. Cell. Probes, 2001, 15, 43-52.
[70] Y. Ishii, Fluorescence resonance energy transfer between single fluorophores attached to a coiled-coil protein in aqueous solution, Chem. Phys. , 247, 1999, 163-173.
[71] R. Yamasakil, Single molecular observation of the interaction of GroEL with substrate proteins, J. Mol. Biol. , 1999, 292, 965-972.
[72] Y. K. Suzukia, Imaging of the fuorescence spectrum of a single fluorescent molecule by prism-based spectroscopy, FEBS Letters, 2002, 512, 235-239.
[73] 杨军,N-十二烷基-N′-(对氨基苯磺酸钠)硫脲的合成,现代化工,1999,19(6),26-27.
[74] 徐克勋,精细有机化工原料及中间体手册,北京:化学工业出版社,3-621.
[75] 马世昌,基础有机化学反应,西安:陕西人民教育出版社,1995,第一版,665.
[76] R. H. Munchen, uber malodialdehyde I, mitteilung, Ber. , 1941, 74 (3), 1825-1829.
[77] S. R Williamt, Two simple amidinium vinylogs, J. Am. Chem. Soc. , 1949, 71, 754-755.
[78] G. L. Stagno, D. Alcontres, G. Lovecchio, Influence of conjugation on the oxidation of alcohols with selenium dioxide, Gazz. Chim. Ital. , 1960, 90, 337-346.
[79] 樊能廷,有机合成事典,北京:北京理工大学出版社,1992,665-666.
[80] M. G. Veliev, M. M. Guseinov, An imprc, ved synthesis of propynal, Synthesis, 1980, 6, 461.
[81] NIST Scientific and Technical Database, http://www.nist.gov/srd/nist35.ntm.
[82] 上海市科学技术交流站组编,正交试验设计法,上海:上海人民出版社,1975.18-23.
[83] B. Tom, H. R. Manchester and H. R. Ernest, United states Patent Office, 1933, 1, 933, 206.
[84] W. H. Warren and F. E. Wilson, Aaction of thionyl chloride on some anilides, ureas and urethane, Ber. 68B, 1935, 957-960.
[85] E. H. John, R. B. Alfred, Unsaturated aromatic amines; a novel synthesis of indoles, J. Am. Chem. Soc. , 1958, 80 (2), 437-439.
[86] P. Karel and G. Hans, A-Halo others, Ⅶ. reactions of dichloromethyl alkyl ethers with amino acid derivatives, Chem. Ber. , 1961, 49, 527-537.
[87] R. Debrath, A. Worth, Symmetrically disubstituted formamidines, formation by raney-nicked rudiction of thioureas, J. Chem. Soc. , 1948, 1716-1720.
[88] M. Passerini, The isonitriles Ⅳ. reaction with organic acids, Gazz. Ber. Ital. , 1922, 52 (Ⅱ), 250-257.
[89] R. M. Roberts, Ortho ester, imilic ester and amidimes, the identity of N-phenyl-N'-p-toylformamidine, J. Am. Chem. Soc. , 72, 3603-3610.
[90] 王晓峰,发明专利申请公开说明书,申请号:89105167.8.公开号CN:1050377A.
[91] 马立人,蒋中华,生物芯片,北京:化学工业出版社,2000,3,11.
[92] 楼开炎,不对称水溶性菁染料的简便制备与纯化方法,华东理工大学学报,2002,28(2),212-215.
[93] W. Harold, A synthesis of bromocaproic acid, J. Am. Chem. Soc. , 1951, 73,