本文選題：金納米顆粒 + 納米電極�。� 參考：《華東師范大學》2015年博士論文

【摘要】：神經(jīng)細胞之間的信息傳遞是通過不同神經(jīng)遞質分子的傳導來實現(xiàn)的。神經(jīng)遞質傳導的紊亂和平衡狀態(tài)的打破是導致多種中樞神經(jīng)系統(tǒng)疾病的重要原因之一。因此,能夠高靈敏、高時空分辨監(jiān)測神經(jīng)遞質在神經(jīng)系統(tǒng)中的動態(tài)釋放,進而研究中樞神經(jīng)系統(tǒng)疾病發(fā)生前后的神經(jīng)遞質水平的變化,將有助于理解中樞神經(jīng)系統(tǒng)障礙或疾病的分子機制,為相關疾病的治療提供方法學指導。 目前,對神經(jīng)遞質進行實時、在體、在線檢測的方法主要有微滲析活體取樣分析技術和在體快速循環(huán)伏安法。微滲析活體采樣可以從生物活體內進行動態(tài)微量生化取樣,有對組織損傷輕、取樣少、可與高效液相、毛細管電泳等分離技術有效結合等優(yōu)點,可應用于實驗動物的所有組織和器官,特別是在神經(jīng)遞質的在體檢測方面的研究和應用,都得到了很好的結果,為探討相關神經(jīng)系統(tǒng)疾病發(fā)病的機制提供了有利的分析方法。而在體快速循環(huán)伏安法由于微電極尺寸小、時空分辨率高、擴散效率和響應速度快等優(yōu)點,目前已經(jīng)被廣泛應用于生物活體分析及單細胞分析等領域。 但是,在體快速循環(huán)伏安法和微滲析活體取樣法只是對擴散至細胞外液中的遞質進行研究,還不能直接檢測肌體受到刺激后突觸間瞬時釋放產(chǎn)生的遞質水平。由于突觸間隙距離很小(小于100nm),若要用電化學方法直接檢測間隙內遞質分子的釋放水平,則需要發(fā)展體積小、靈敏度高、時空分辨率高、直徑小于100nm的納米電極。為了制備適于突觸間隙分析的納米電極,本論文設計了簡單、綠色環(huán)保的方法制備了尺寸可控的金納米電極。在此基礎上,通過在納米電極組裝特異性分子,發(fā)展了具有良好選擇性、穩(wěn)定性、靈敏度和生物相容性的功能納米電極,實現(xiàn)對神經(jīng)遞質的高選擇性的直接檢出。全文共分為五個部分,具體內容如下： 第一章緒論 本章主要介紹了納米電極的研究背景、納米電極的研究意義、納米電極的分類及制備方法以及納米電極在生物化學中的應用,著重介紹了納米電極的制備方法及其表征和納米電極的應用。 第二章基于混合包裹法和電化學刻蝕可控制備盤狀金納米電極 本工作中,我們先利用電化學刻蝕然后采用混合包裹法制備了一種新型的尺寸可控的盤狀金納米電極,該方法簡單、綠色環(huán)保。通過TEM和穩(wěn)態(tài)循環(huán)伏安法對盤狀金納米電極尖端進行了表征,結果表明：金納米尖端最小半徑為～6nm；電極外部混合絕緣層薄而均勻,厚度在30m左右,經(jīng)過包裹后的電極尖端整體大小仍能保持在納米級別；在KCl溶液里通過循環(huán)伏安掃描刻蝕裸露的金尖端后得到盤狀金納米電極。電極在K3Fe(CN)6、Fc、K3IrCl6三種標準體系中進行穩(wěn)態(tài)循環(huán)伏安掃描,循環(huán)伏安曲線具有很好的“S”形貌,說明該方法制備的電極具備了納米電極的特征；并利用所得的穩(wěn)態(tài)極限電流準確地求出電極的有效半徑；電極的尺寸可控且電極的制備重現(xiàn)性好,電極表面的絕緣層在水相和有機相體系中均表現(xiàn)出了良好的穩(wěn)定性。該部分工作為后續(xù)動力學參數(shù)的求算和生物中的應用提供了有利的實驗工具。 第三章盤狀金納米電極的電化學行為及其在單細胞檢測中的應用 本工作中,通過穩(wěn)態(tài)循環(huán)伏安法,利用不同有效半徑的盤狀金納米電極在K3Fe(CN)6、Fc和K3IrCl6三種標準體系中進行穩(wěn)態(tài)循環(huán)伏安掃描,研究盤狀金納米電極表面的動力學行為,通過統(tǒng)計數(shù)據(jù)求出了三個體系中電極表面的傳遞系數(shù)(a)和異相電子轉移速率常數(shù)(k°),實驗結果與文獻報道一致,表明盤狀納米電極表面物質的傳遞速率快,具有較大的異相電子轉移速率常數(shù)等優(yōu)越的電化學性能,有利于電極在生物領域中的應用。電極經(jīng)nafion修飾后,對多巴胺具有較好的選擇性、靈敏度高、低的檢測限和寬的線性范圍等優(yōu)點。單細胞實驗中,選用PC12(腎上腺嗜鉻細胞瘤)細胞為研究對象,nafion修飾的盤狀金納米電極成功地監(jiān)測到了PC12單細胞中的單個囊泡中的胞吐釋放,PC12單細胞中囊泡內的主要成分為多巴胺,與其他文獻報道的類似。單細胞膜上并不是所有地方都有囊泡釋放,釋放點在細胞上成散點分布；活性區(qū)域內,在一個釋放點釋放的囊泡個數(shù)也無明顯規(guī)律；即使是非常相近的活性釋放點,釋放的囊泡個數(shù)也不相同。說明本工作中所制備的納米電極靈敏度高、檢測限低、生物相容性較好,為后續(xù)檢測神經(jīng)細胞突觸間神經(jīng)遞質分子的釋放提供了潛在的分析工具。 第四章納米毛細管尖端外壁化學沉積金納米顆粒制備金納米電極及其對鼠腦中多巴胺的分析檢測 本工作中,主要論述在石英毛細管錐形尖端外壁附著一層由金納米粒子構成的金膜,通過控制電泳時間、電泳電位、電泳次數(shù)、金納米顆粒的沉積時間等來制備一種尺寸可控的金納米電極,及其經(jīng)nafion修飾后在活體分析中的應用。首先,通過調節(jié)激光拉制儀的五個參數(shù)將石英毛細管拉制成納米尺寸的錐形尖端；然后依次經(jīng)過羥基化、硅烷偶聯(lián)劑修飾、附著金納米顆粒生長因子、在氯金酸和鹽酸羥胺混合生長液中附著由金納米顆粒形成的金膜；最后將附著金納米顆粒的錐形尖端制備成金電極,經(jīng)陰極電泳漆絕緣包裹即可以得到金納米電極。SEM結果表明電極尖端形貌符合半球狀金納米電極的特征。電化學實驗結果表明：電極在鐵氰化鉀和三氯化六氨合釕標準溶液中均呈現(xiàn)良好的“S”圖譜,經(jīng)計算得出該類電極尺寸大小達到了納米級別。經(jīng)Nafion修飾的金納米電極對多巴胺有良好的選擇性,線性范圍寬,在5.6×10～52×10-8M之間均呈現(xiàn)了良好的線性響應,靈敏度高、檢測限低(1×10-8M)。該工作所制備的金納米電極除尺寸小等優(yōu)點外,以石英光纖為基底大大地提高了電極的韌性和硬度及電極壽命,成功地被植入已麻醉的SD大鼠大腦紋狀體區(qū)域并得到了在線的電流-電位曲線,結果表明老鼠紋狀體內多巴胺濃度可受外部藥物刺激的影響而增大,濃度最高可達49nM。為后續(xù)在鼠腦等生物組織中直接檢測神經(jīng)遞質分子提供了新的分析方法和工具。 第五章不同形貌的金納米材料在金納米電極表面的電化學行為研究 本工作中,我們利用激光拉制法和精細打磨的方法,成功地制備出了尺寸可控、穩(wěn)定性好的金納米電極,并以此為基底分別附著上不同形貌的金納米粒子以考察不同形貌的金納米粒子對多巴胺的響應。借助光學顯微鏡、SEM, TEM對電極的表面形貌進行了表征。通過穩(wěn)態(tài)循環(huán)伏安法計算電極的有效半徑其最小可達6.82nm。在+200mV下,經(jīng)nafion修飾的電極尖端附著了不同形貌的金納米顆粒的金納米電極對多巴胺都有良好的響應,其中錐形金納米顆粒本身因具有較好的自凈能力和很好的穩(wěn)定性,對多巴胺的響應最好。經(jīng)校準,對多巴胺的檢測限(S/N=3)分別為：錐形金納米顆粒(AuNP):～5.2×10-9M,棒形金納米顆粒(AuNR):～1.71×10-8M,球形金納米顆粒(AuNS):～3.2×10-8M。錐形金納米顆粒對多巴胺的線性范圍為：1×10-8～2.55×10-6M,該類電極具有良好的重現(xiàn)性、穩(wěn)定性,且電極的利用率很高。以附著有錐形納米顆粒的金納米電極為工作電極植入已麻醉的大鼠紋狀體內在線研究了腦內多巴胺的釋放水平。
[Abstract]:The transmission of information between nerve cells is achieved through the conduction of different neurotransmitters . The disorder of neurotransmitter conduction and the breaking of equilibrium state are one of the important causes of various central nervous system diseases . Therefore , it is possible to monitor the dynamic release of neurotransmitters in the nervous system with high sensitivity and high space - time resolution , and then to study the changes of the neurotransmitter levels before and after the onset of the central nervous system disease , which will help to understand the molecular mechanism of the disorders or diseases of the central nervous system and provide methodological guidance for the treatment of related diseases .

The present invention has the advantages of low micro - electrode size , high space - time resolution , high diffusion efficiency , high response speed and the like , and is widely applied to the fields of living body analysis and single - cell analysis .

In order to prepare the nanoelectrodes suitable for synaptic cleft analysis , the nanoelectrodes with controllable size can be directly detected by electrochemical methods . In order to prepare the nano electrodes suitable for synaptic cleft analysis , the method is simple and environmentally friendly . In order to prepare the nano electrodes suitable for synaptic cleft analysis , the direct detection of high selectivity of neurotransmitters is realized .

Chapter 1 Introduction

This chapter mainly introduces the research background of nano - electrode , the research significance of nano - electrode , the classification of nano - electrode , its preparation method and the application of nano - electrode in biochemistry , focusing on the preparation method and characterization of nano - electrode and the application of nano - electrode .

The second chapter is based on hybrid wrapping method and electrochemical etching to control the disk - shaped gold nano - electrode .

In this work , a novel size - controllable disc - shaped gold electrode with controllable size is prepared by electrochemical etching . The method is simple , green and environment - friendly , and the tip of the disk - shaped gold electrode is characterized by TEM and steady - state cyclic voltammetry . The results show that the minimum radius of the gold nanometer tip is ~ 6 nm ;
the external mixed insulating layer of the electrode is thin and uniform , the thickness is about 30 m , and the overall size of the electrode tip after the wrapping can be kept at the nanometer level ;
The disk - shaped gold nano - electrodes were obtained by cyclic voltammetry scanning in KCl solution . The electrodes were subjected to steady - state cyclic voltammetry scanning in three standard systems of K _ 3Fe ( CN ) 6 , Fc and K _ 3 IrCl6 , and the cyclic voltammogram has a good " S " shape , and the electrode prepared by the method has the characteristics of nano - electrode .
and using the obtained steady - state limit current to accurately calculate the effective radius of the electrode ;
the size of the electrode is controllable and the preparation reproducibility of the electrode is good , and the insulating layer on the surface of the electrode shows good stability in the water phase and the organic phase system .

The Electrochemical Behavior and Its Application in Single Cell Detection

In this work , by steady - state cyclic voltammetry , the dynamic behavior of the surface of the disk - shaped gold nano - electrode was investigated by means of steady - state cyclic voltammetry scanning in three standard systems of K _ 3Fe ( CN ) 6 , Fc and K _ 3 IrCl6 .
There was no obvious regularity in the number of vesicles released at one release point in the active area .
The prepared nano electrode is high in sensitivity , low in detection limit and good in biocompatibility , and provides a potential analysis tool for subsequent detection of the release of nerve cell synaptic neurotransmitter molecules .

Preparation of gold nanoparticles by chemical deposition of gold nanoparticles on the outer wall of nano capillary tip and analysis of dopamine in rat brain

In this work , a gold film composed of gold nanoparticles is attached to the outer wall of the conical tip of the quartz capillary , and the gold nano - electrode with controllable size is prepared by controlling the electrophoresis time , the electrophoresis potential , the number of electrophoresis , the deposition time of the gold nanoparticles and the like .
then the gold film formed by gold nanoparticles is attached in the mixed growth liquid of chloroauric acid and hydroxylamine hydrochloride through hydroxylation , silane coupling agent modification and gold nanoparticle growth factor ;
The results show that the electrode has a good linear response to dopamine in the standard solution of potassium cyanide and ruthenium trichloride . The results show that the electrode has a good selectivity to dopamine and has high sensitivity and low detection limit ( 1 脳 10 - 8M ) . The results show that the concentration of dopamine in the striatum of rat striatum can be increased by the influence of external drug stimulation . The results show that the concentration of dopamine in striatum of rats is up to 49 nM .

Study on the Electrochemical Behavior of Gold Nanomaterials with Different Morphology on the Surface of Gold Nanoelectrodes

In this work , we have successfully prepared gold nano - particles with controllable size and good stability by laser - drawing and fine - polishing . The surface features of the electrodes are characterized by optical microscopy , SEM and TEM . The linear range of the electrodes is 1 脳 10 - 8 锝，

本文編號：1883966