本文選題：縮膽囊素 + 腦啡肽　； 參考：《武漢大學(xué)》2017年碩士論文

【摘要】：縮膽囊素神經(jīng)肽和腦啡肽是廣泛分布于中樞神經(jīng)系統(tǒng)中的兩類多功能性神經(jīng)肽,對(duì)多種生理過(guò)程如神經(jīng)傳遞,癲癇,抑郁,痛覺(jué)等有重要的調(diào)控作用。目前報(bào)道的腦脊液及血液中縮膽囊素神經(jīng)肽和腦啡肽的定量主要基于放射免疫法和酶聯(lián)免疫法,這些免疫分析方法雖然靈敏度高,但需使用昂貴的抗體,且操作繁瑣,存在輻射污染等問(wèn)題。因此,研發(fā)經(jīng)濟(jì)、高特異性的人工抗體來(lái)代替生物抗體用于多肽及蛋白質(zhì)的分析檢測(cè)具有重大意義。分子印跡聚合物生物芯片是將分子印跡技術(shù)和生物芯片相結(jié)合,對(duì)目標(biāo)分子具有專一性識(shí)別能力的新型生物芯片,兼具分子印跡聚合物高特異性,耐酸堿,抗高溫和生物芯片高通量,快速,微量消耗的特點(diǎn)。本研究將分子印跡聚合物的高選擇性識(shí)別能力、微陣列芯片和微孔板芯片的高通量以及熒光檢測(cè)的高靈敏度三大優(yōu)勢(shì)相結(jié)合,研究制備腦啡肽分子印跡聚合物陣列芯片、縮膽囊素分子印跡聚合物微孔板芯片和縮膽囊素分子印跡膜,基于此陣列芯片和微孔板芯片建立了熒光免疫分析方法對(duì)人腦脊液中低豐度的縮膽囊素神經(jīng)肽和腦啡肽進(jìn)行定量分析。本文主要進(jìn)行了以下研究:1.研究制備腦啡肽分子印跡聚合物微陣列芯片。將光刻技術(shù)與抗原決定簇法相結(jié)合,通過(guò)光刻掩膜法在硅烷化的玻片表面原位聚合制備對(duì)腦啡肽具有特異性識(shí)別能力的分子印跡聚合物微陣列芯片。研究并優(yōu)化印跡聚合物陣列芯片合成制備條件。借助分子模擬,計(jì)算模板-單體復(fù)合物之間的結(jié)合能;用掃描電鏡和紅外光譜等對(duì)印跡材料進(jìn)行表征。以平衡吸附實(shí)驗(yàn)和MALDI-TOF-MS分析研究評(píng)估印跡材料特異性及專一性。最后基于該印跡聚合物芯片,研究建立了固相競(jìng)爭(zhēng)熒光免疫分析方法,利用倒置熒光顯微鏡—CCD圖像分析系統(tǒng)對(duì)人腦脊液中低豐度腦啡肽進(jìn)行定量分析。2.縮膽囊素神經(jīng)肽分子印跡聚合物微孔板芯片的制備及熒光免疫分析應(yīng)用。采用抗原決定簇法沉淀聚合制備對(duì)縮膽囊素神經(jīng)肽具有特異性識(shí)別能力的分子印跡聚合物微球,以此MIP微球代替昂貴的縮膽囊素神經(jīng)肽抗體,用聚乙烯醇(PVA)固定于96孔板中,制備縮膽囊素神經(jīng)肽分子印跡聚合物微孔板芯片。優(yōu)化印跡聚合物微孔板芯片的合成制備條件,用掃描電鏡對(duì)印跡微球進(jìn)行表征。以平衡吸附實(shí)驗(yàn)探索印跡材料特異性及選擇性。最后基于分子印跡聚合物微孔板芯片,研究建立固相競(jìng)爭(zhēng)熒光免疫分析方法,利用倒置熒光顯微鏡—CCD圖像分析系統(tǒng)對(duì)人腦脊液中縮膽囊素神經(jīng)肽進(jìn)行定量分析。3.縮膽囊素神經(jīng)肽分子印跡聚合膜的制備及其熒光免疫分析應(yīng)用。采用抗原決定簇法,在硅烷化的玻片表面原位合成制備對(duì)縮膽囊素神經(jīng)肽具有特異性識(shí)別能力的分子印跡膜。優(yōu)化印跡膜合成制備條件,并用掃描電鏡和紅外光譜對(duì)其進(jìn)行表征,通過(guò)吸附平衡實(shí)驗(yàn)對(duì)印跡膜的吸附容量及選擇性進(jìn)行評(píng)價(jià)。最后基于該分子印跡膜,研究建立固相競(jìng)爭(zhēng)熒光免疫分析方法,利用倒置熒光顯微鏡—CCD圖像分析系統(tǒng)對(duì)人腦脊液中縮膽囊素神經(jīng)肽進(jìn)行定量分析。
[Abstract]:Cholecystokinin and enkephalin are two kinds of multifunctional neuropeptides widely distributed in the central nervous system. They have important regulatory effects on many physiological processes, such as neurotransmission, epilepsy, depression, and pain. The current reported quantitative of cholecystokinin and enkephalin in cerebrospinal fluid and blood is mainly based on radioimmunoassay and enzyme. Although these immunoassays are sensitive, they need to use expensive antibodies and have complicated operation and radiation pollution. Therefore, it is of great significance to develop economic, highly specific artificial antibodies instead of biological antibodies for the analysis and detection of peptides and proteins. Molecular imprinted polymer biochips are the molecules of the molecular imprinted polymer. A new biochip with the ability to identify the target molecules with the specificity of the molecularly imprinted polymer, with the characteristics of high specificity of molecularly imprinted polymers, acid resistance and alkali resistance, high temperature resistance and high throughput, rapid and micro consumption of biochips. The high selectivity recognition ability of molecularly imprinted polymers, microarray chips and micropores in this study The high throughput of the chip and the high sensitivity of the fluorescence detection are combined to study the preparation of the enkephalin molecularly imprinted polymer array chip, the molecular imprinted polymer microchip and the cholecystokinin molecularly imprinted membrane. Based on this array chip and microplate chip, the fluorescence immunoassay method is established for the human cerebrospinal fluid (CSF). The quantitative analysis of the moderate and low abundance of cholecystokinin neuropeptides and enkephalin was carried out. The following studies were carried out in this paper: 1. the preparation of enkephalin molecularly imprinted polymer microarray was studied. The photolithography technique was combined with the antigen determinant method to prepare enkephalin on the surface of silanized glass by photolithography. Molecularly imprinted polymer microarray for sexual recognition. Study and optimize the synthesis conditions of imprinted polymer array chips. By means of molecular simulation, the binding energy between template and monomers is calculated. The imprinted materials are characterized by scanning electron microscopy and infrared spectroscopy. The evaluation of the adsorption and MALDI-TOF-MS analysis is evaluated by the scanning electron microscopy and infrared spectroscopy. The specificity and specificity of the imprinted material. Finally, based on the imprinted polymer chip, the solid phase competitive fluorescence immunoassay was established, and the quantitative analysis of the low abundance enkephalin in the human cerebrospinal fluid by the inverted fluorescence microscope CCD image analysis system was used to prepare the.2. cholecystokinin neuropeptide molecularly imprinted polymer microplate chip The molecularly imprinted polymer microspheres with specific recognition ability for cholecystokinin neuropeptides were prepared by precipitation polymerization with antigenic determinant method. The MIP microspheres were replaced by the expensive cholecystokinin neuropeptide antibody and polyvinyl alcohol (PVA) was immobilized in 96 pore plates to prepare the cholecystokinin neuropeptide molecularly imprinted polymer. Microporous plate chip. Optimization of synthesis conditions of imprinted polymer microplate chips, characterization of imprinted microspheres by scanning electron microscopy. The specificity and selectivity of imprinted materials are explored by equilibrium adsorption experiments. Finally, based on molecularly imprinted polymer microplate chips, solid phase competitive fluorescence immunoassay method was established and inverted fluorescence was used. Quantitative analysis of cholecystokinin neuropeptide in human cerebrospinal fluid by micromirror CCD image analysis system the preparation of.3. cholecystokinin neuropeptide molecularly imprinted polymer membrane and its application in fluorescence immunoassay. The preparation conditions of the imprinted membrane were optimized, and the preparation conditions were synthesized and characterized by scanning electron microscope and infrared spectrum. The adsorption capacity and selectivity of the imprinted membrane were evaluated by adsorption equilibrium experiment. Finally, based on the molecularly imprinted membrane, the solid phase competitive fluorescence immunoassay method was established, and the inverted fluorescence microscope CCD image was used. A systematic analysis of cholecystokinin neuropeptides in human cerebrospinal fluid was carried out.

【學(xué)位授予單位】：武漢大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：O631.3;O657.3

【參考文獻(xiàn)】

相關(guān)期刊論文 前2條

1 Zhong-Shuai Wu;Xinliang Feng;Hui-Ming Cheng;;Recent advances in graphene-based planar micro-supercapacitors for on-chip energy storage[J];National Science Review;2014年02期

2 衛(wèi)瀟;潘建明;戴江棟;閆永勝;;表面分子印跡技術(shù)在分析化學(xué)中的應(yīng)用研究進(jìn)展[J];離子交換與吸附;2013年02期

，

本文編號(hào)：1841514