酞菁紅區(qū)熒光探針在生物大分子檢測中的應(yīng)用及其用于生物成像的可行性初探
發(fā)布時間：2018-11-13 15:39
【摘要】：由于在長波區(qū)域具有吸收或發(fā)射特性的天然和人工合成的物質(zhì)極少,因而在進(jìn)行生物、生化和臨床等復(fù)雜試樣的測定時,使用長波發(fā)射熒光試劑可有效避開背景熒光和散射光的干擾,且光漂白作用小,與傳統(tǒng)的熒光試劑相比具有很大優(yōu)越性。能發(fā)熒光的酞菁化合物即是一類紅區(qū)發(fā)射的熒光試劑。本論文的工作圍繞酞菁紅區(qū)熒光探針在生物大分子分析及成像檢測中的應(yīng)用而展開,共分五章。 第一章：首先對紅區(qū)及近紅外熒光探針的應(yīng)用進(jìn)展進(jìn)行了簡要介紹。由于酞菁化合物作為紅區(qū)熒光試劑的研究構(gòu)成了本文的主要部分,本章還對酞菁化合物的分子結(jié)構(gòu)和光譜特性以及它在生物模擬酶、光動力治療、生化分析領(lǐng)域的應(yīng)用做了介紹。 第二章：本章建立了簡便、快速測定硫酸軟骨素的熒光增強(qiáng)分析法。通過熒光光譜篩選實驗發(fā)現(xiàn),具有共軛結(jié)構(gòu)的陽離子頭部和長碳鏈尾部的陽離子表面活性劑可幾乎完全猝滅四磺基鋁酞菁(Tetrasulfonated Aluminum Phthalocyanine, AlS4Pc)的熒光。而在帶有磺基陰離子的硫酸軟骨素(Chondroitin Sulfate, CS)存在下,體系熒光顯著恢復(fù)。以AIS4Pc-陽離子表面活性劑離子締合物為紅區(qū)熒光探針,考察了其對CS的熒光恢復(fù)響應(yīng)行為,發(fā)現(xiàn)離子締合物的熒光恢復(fù)程度與CS濃度存在良好的相關(guān)性,據(jù)此實現(xiàn)了復(fù)雜樣品中CS簡便、準(zhǔn)確的測定。本章工作分為兩部分：由于前期工作篩選出的猝滅效果較好的陽離子表面活性劑有兩種,由這兩種猝滅劑參與的體系的靈敏度和線性范圍不同,各具特色,因此對兩個體系分別進(jìn)行了考察,并建立了兩種CS的熒光增強(qiáng)測定法。 第三章：本章建立了簡便、快速測定溶菌酶的熒光增強(qiáng)分析法。帶有強(qiáng)陰離子的黏多糖肝素(Heparin, HP)可誘導(dǎo)陽離子鋁酞菁[Tetra(trimethyammionio) Aluminum Phthalocynine, TTMAAlPc]聚集而導(dǎo)致熒光顯著猝滅。由于溶菌酶對黏多糖具有催化降解作用,因而可水解HP為小分子片段,從而破壞TTMAAlPc-HP的聚集締合平衡,使TTMAAlPc被釋放,體系熒光將因之恢復(fù)。結(jié)合熒光光譜與熒光各向異性技術(shù)對反應(yīng)機(jī)理進(jìn)行了探討。據(jù)此建立了溶菌酶測定新方法,并實現(xiàn)了復(fù)雜樣品中的溶菌酶含量的準(zhǔn)確測定。 第四章：探討了紅區(qū)熒光染料酞菁在指紋成像觀測中的應(yīng)用�？疾炝藥追N酞菁染料對潛在指印的成像效果,其中A1(SO2C1)4TSP對于玻片上油指印的染色效果最為理想,進(jìn)一步的對染料溶劑,濃度,染色方法和時間進(jìn)行了考察和優(yōu)化,成像結(jié)果令人滿意。 第五章：為研究紅區(qū)熒光染料酞菁在細(xì)胞中是否有特異性定位,為此首先通過摸索建立了顯微注射技術(shù),直接將熒光酞菁染料注射進(jìn)細(xì)胞中,體外培養(yǎng)后觀察染料與細(xì)胞是否有結(jié)合。初步研究的結(jié)果表明熒光酞菁化合物具有成為活細(xì)胞成像新型熒光探針的潛力。
[Abstract]:Since there are very few natural and synthetic substances with absorption or emission characteristics in the long-wave region, when determining complex samples, such as biological, biochemical and clinical, The use of long wave emission fluorescence reagent can effectively avoid the interference of background fluorescence and scattered light, and the photobleaching effect is small, which is superior to the traditional fluorescent reagent. The fluorescent phthalocyanine compounds are a kind of fluorescent reagents in the red region. This paper focuses on the application of phthalocyanine red region fluorescence probe in biomolecules analysis and imaging detection, which is divided into five chapters. Chapter 1: firstly, the application progress of red region and near infrared fluorescence probe is briefly introduced. Since the study of phthalocyanine compounds as a red region fluorescent reagent constitutes the main part of this paper, this chapter also deals with the molecular structure and spectral characteristics of phthalocyanine compounds, as well as its biological mimic enzyme, photodynamic therapy, The application of biochemical analysis is introduced. Chapter 2: in this chapter, a simple and rapid fluorescence enhanced method for the determination of chondroitin sulfate was established. The fluorescence spectra showed that the cationic surfactants with conjugated structure and long carbon chain tail could almost completely quench the fluorescence of the tetrasulfonyl aluminum phthalocyanine (Tetrasulfonated Aluminum Phthalocyanine, AlS4Pc). In the presence of chondroitin sulfate (Chondroitin Sulfate, CS) with sulfonyl anion, the fluorescence of the system recovered significantly. Using AIS4Pc- cationic surfactant ion-association complex as a red region fluorescence probe, the fluorescence recovery response to CS was investigated. It was found that there was a good correlation between the fluorescence recovery degree of the ion-association complex and the concentration of CS. Based on this, the determination of CS in complex samples was simple and accurate. The work of this chapter is divided into two parts: because there are two kinds of cationic surfactants with better quenching effect, the sensitivity and linear range of the two quenching agents are different and have their own characteristics. Therefore, the two systems were investigated, and two methods of fluorescence enhanced determination of CS were established. Chapter 3: in this chapter, a simple and rapid fluorescence enhanced assay for the determination of lysozyme was established. The strong anionic mucopolysaccharide heparin (Heparin, HP) can induce cationic aluminum phthalocyanine [Tetra (trimethyammionio) Aluminum Phthalocynine, TTMAAlPc] aggregation and lead to fluorescence quenching. Because lysozyme can catalyze the degradation of mucopolysaccharide, HP can be hydrolyzed as a small molecular fragment, thus destroying the aggregation and association equilibrium of TTMAAlPc-HP, releasing TTMAAlPc and restoring the fluorescence of the system. The mechanism of the reaction was discussed by means of fluorescence spectroscopy and fluorescence anisotropy. A new method for the determination of lysozyme was established, and the accurate determination of lysozyme in complex samples was achieved. Chapter 4: the application of fluorescent dye phthalocyanine in fingerprint imaging is discussed. The imaging effects of several phthalocyanine dyes on potential fingerprinting were investigated. The dyeing effect of A1 (SO2C1) 4TSP on oil fingerprinting on glass was the most ideal. The solvent, concentration, dyeing method and time of dye were further investigated and optimized. The imaging results are satisfactory. Chapter 5: in order to study whether the red zone fluorescent dye phthalocyanine has a specific localization in the cell, a microinjection technique was established to directly inject the fluorescent phthalocyanine dye into the cell. After in vitro culture, the binding of dye to cells was observed. The preliminary results show that fluorescent phthalocyanines have the potential to be novel fluorescent probes for living cell imaging.
【學(xué)位授予單位】：廈門大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R927;O657.3
本文編號：2329578
[Abstract]:Since there are very few natural and synthetic substances with absorption or emission characteristics in the long-wave region, when determining complex samples, such as biological, biochemical and clinical, The use of long wave emission fluorescence reagent can effectively avoid the interference of background fluorescence and scattered light, and the photobleaching effect is small, which is superior to the traditional fluorescent reagent. The fluorescent phthalocyanine compounds are a kind of fluorescent reagents in the red region. This paper focuses on the application of phthalocyanine red region fluorescence probe in biomolecules analysis and imaging detection, which is divided into five chapters. Chapter 1: firstly, the application progress of red region and near infrared fluorescence probe is briefly introduced. Since the study of phthalocyanine compounds as a red region fluorescent reagent constitutes the main part of this paper, this chapter also deals with the molecular structure and spectral characteristics of phthalocyanine compounds, as well as its biological mimic enzyme, photodynamic therapy, The application of biochemical analysis is introduced. Chapter 2: in this chapter, a simple and rapid fluorescence enhanced method for the determination of chondroitin sulfate was established. The fluorescence spectra showed that the cationic surfactants with conjugated structure and long carbon chain tail could almost completely quench the fluorescence of the tetrasulfonyl aluminum phthalocyanine (Tetrasulfonated Aluminum Phthalocyanine, AlS4Pc). In the presence of chondroitin sulfate (Chondroitin Sulfate, CS) with sulfonyl anion, the fluorescence of the system recovered significantly. Using AIS4Pc- cationic surfactant ion-association complex as a red region fluorescence probe, the fluorescence recovery response to CS was investigated. It was found that there was a good correlation between the fluorescence recovery degree of the ion-association complex and the concentration of CS. Based on this, the determination of CS in complex samples was simple and accurate. The work of this chapter is divided into two parts: because there are two kinds of cationic surfactants with better quenching effect, the sensitivity and linear range of the two quenching agents are different and have their own characteristics. Therefore, the two systems were investigated, and two methods of fluorescence enhanced determination of CS were established. Chapter 3: in this chapter, a simple and rapid fluorescence enhanced assay for the determination of lysozyme was established. The strong anionic mucopolysaccharide heparin (Heparin, HP) can induce cationic aluminum phthalocyanine [Tetra (trimethyammionio) Aluminum Phthalocynine, TTMAAlPc] aggregation and lead to fluorescence quenching. Because lysozyme can catalyze the degradation of mucopolysaccharide, HP can be hydrolyzed as a small molecular fragment, thus destroying the aggregation and association equilibrium of TTMAAlPc-HP, releasing TTMAAlPc and restoring the fluorescence of the system. The mechanism of the reaction was discussed by means of fluorescence spectroscopy and fluorescence anisotropy. A new method for the determination of lysozyme was established, and the accurate determination of lysozyme in complex samples was achieved. Chapter 4: the application of fluorescent dye phthalocyanine in fingerprint imaging is discussed. The imaging effects of several phthalocyanine dyes on potential fingerprinting were investigated. The dyeing effect of A1 (SO2C1) 4TSP on oil fingerprinting on glass was the most ideal. The solvent, concentration, dyeing method and time of dye were further investigated and optimized. The imaging results are satisfactory. Chapter 5: in order to study whether the red zone fluorescent dye phthalocyanine has a specific localization in the cell, a microinjection technique was established to directly inject the fluorescent phthalocyanine dye into the cell. After in vitro culture, the binding of dye to cells was observed. The preliminary results show that fluorescent phthalocyanines have the potential to be novel fluorescent probes for living cell imaging.
【學(xué)位授予單位】：廈門大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R927;O657.3
【參考文獻(xiàn)】
相關(guān)期刊論文 前10條
1 周葉;趙婷;馮偉偉;李蕓子;但招陵;劉治華;仰榴青;;咔唑法測定中華鱘軟骨中硫酸軟骨素的含量[J];安徽農(nóng)業(yè)科學(xué);2011年19期
2 余遠(yuǎn)斌,楊錦宗;酞菁類催化劑的研究進(jìn)展[J];北京工業(yè)大學(xué)學(xué)報;1998年02期
3 呂義,李景舜,王春華,隋春生,王渭;城鄉(xiāng)學(xué)齡兒童唾液溶菌酶含量正常值的探討[J];白求恩醫(yī)科大學(xué)學(xué)報;1993年06期
4 馬春艷;付曉燕;張藝娟;楊溢;牛淑云;金晶;張洪武;辛勤;;納米近紅外熒光技術(shù)在活體成像上的應(yīng)用[J];材料導(dǎo)報;2011年S2期
5 李東輝,楊黃浩,鄭洪,陳賓,陳秋影,許金鉤;用四磺基鋁酞菁直接測定血清中白蛋白[J];分析化學(xué);1999年09期
6 魏玲,楊黃浩,陳小蘭,曲會英,許金鉤;四磺基鋁酞菁與愛爾新藍(lán)的締合作用在核酸定量中的應(yīng)用[J];分析化學(xué);2002年08期
7 李永新,趙丹華,卓淑娟,朱昌青,王倫;四磺基錳酞菁作為過氧化物模擬酶在過氧化氫測定中的應(yīng)用[J];分析化學(xué);2003年06期
8 劉保生,高靜,楊更亮;吖啶橙羅丹明6G熒光共振能量轉(zhuǎn)移及其羅丹明6G熒光猝滅法測定蛋白質(zhì)[J];分析化學(xué);2005年04期
9 胡敏;張鎮(zhèn)西;沈國勵;劉婭莉;;近紅外熒光探針天青A檢測DNA[J];分析化學(xué);2007年06期
10 王峧胤;王德艷;汪國秀;胡效亞;;基于原子力顯微鏡的微懸臂傳感器測定溶菌酶[J];分析化學(xué);2010年12期
，本文編號：2329578
本文鏈接：http://sikaile.net/yixuelunwen/yiyaoxuelunwen/2329578.html
最近更新
教材專著
