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研究生: 張宏維
Hung-wai Chang
論文名稱: 嗎啡免疫生化感測器研究與應用
Preparation and application of morphine Immunosensors
指導教授: 施正雄
Shih, Jeng-Shong
學位類別: 碩士
Master
系所名稱: 化學系
Department of Chemistry
論文出版年: 2002
畢業學年度: 90
語文別: 中文
論文頁數: 100
中文關鍵詞: 嗎啡
英文關鍵詞: morphine
論文種類: 學術論文
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  • 目前在檢測血液或尿液中嗎啡量常用的方法是利用管柱層析如氣象層析儀和高效能液相層析儀兩種方法。但是這樣的方法中往往需要事前的樣品處理需要相當多的時間與人力,免疫方法可以解決管柱耗時的問題。本篇所用的方法是利用抗嗎啡抗體與標幟上嗎啡的G6PDH(Glucose-6-phosphate dehydrogenase labeled with morphine, G6PDH-morphine)在光學與石英壓電免疫感測器上來進行偵測。在免疫感測器的部分,有很好的線性關係,而且偵測下限是113 ng/ml,這樣的偵測下限遠低於目前法律規定的300ng/ml,因此可以應用在實際的檢驗上,且光學免疫感測器的相對標準偏差值(R.S.D.) 3%左右,這是不錯的再線性的檢測方法。並且本篇也探討體內血清與尿液中主要的成分對於這個感測方法的影響。在石英壓電免疫感測器上,首先製備C60-G6PDH並且把C60-G6PDH固定在石英壓電晶體上作為感測膜來對於溶液中抗嗎啡抗體,當C60-G6PDH的吸附溶液中抗嗎啡抗體後會造成頻率的下降來作為偵測的標準,利用固定C60-G6PDH的石英壓電晶體來偵測溶液中的抗嗎啡抗體有相當不錯的再現性且對於抗嗎啡抗體有不錯的選擇性。

    Current technology for determination of morphine in blood or urine is often to use the chromatographic methods such as GC and HPLC, but most of these methods are time consuming. To solve this problem, a morphine optical immuno-sensor and a quartz piezoelectric crystal sensor based on anti-morphine antibody and Glucose-6-phosphate dehydrogenase labeled with morphine (G6PDH-morphine) for opiate solutions were designed and prepared in this study. The optical morphine sensor exhibited good sensitivity with a detection limit of 113 ng/mL which was far below the allowed concentration (300 ng/mL) in law for morphine in biological solutions. The optical sensor also showed a high reproducibility with a relative standard deviation (RSD) of approximately 3 %. The interference of major biological species in simulated serum and urine solutions for the determination of morphine with the optical immuno-sensor was investigated and discussed. In the preparation of the quartz piezoelectric sensor, immobilized fullerene C60 - G6PDH-morphine was synthesized and applied as a coating material on quartz piezoelectric crystals for the immunoassay of anti-morphine antibody in aqueous solutions. The oscillating frequency of the G6PDH-morphine coated-piezoelectric crystal of the piezoelectric sensor decreased by adsorption of anti-morphine antibody in solutions. The piezoelectric crystal sensor showed nearly no interference from various biological species in simulated serum. The piezoelectric crystal sensor based on immobilized C60-G6PDH-morphine for anti-morphine antibody also exhibited good sensitivity and good reproducibility.

    英文摘要………………………………………………………………...I 中文摘要……………………………………………………………...II 目錄…………………………………………………………………..III 圖目錄…………………………………………………………….….III 表目錄……………………………...………………………………VIII 第一章 緒論……………………………………………………………1 1-1 嗎啡簡介………………………………………………………...1 1-1-1 藥物濫用………………………………………………………1 1-1-2 毒品種類的區分………………………………………………2 1-1-3 鴉片、嗎啡、海洛因簡介……………………………………3 1-1-4 嗎啡的藥理作用………………………………………………5 1-1-5嗎啡常用的分析方法………………………………………….9 1-1-6 法定嗎啡的分析方法………………………………………...13 1-2 免疫分析…………………………………………………………..13 1-2-1放射免疫分析法………………………………………………..14 1-2-2 螢光免疫分析法……………………………………………….15 1-2-3 酵素免疫分析法……………………………………………….15 1-2-3-1酵素連結免疫吸附法ELISA(Enzyme-link immunosorbent assay…………...…………………………………………………..16 (一)三明治型(sandwich technique)……………………………….16 (二)競爭型(competitive technique)………………………….17 1-2-3-2酵素增幅免疫分析法EMIT(Enzyme Multiplied Immunoassay Technique)…………………………………………..………….….19 1-3 免疫感測器(Immunosasensors)…………………………..….23 1-3-1 壓電免疫感測器(Piezoelectric immunosasensors)…….24 1-3-2 光學免疫感測器(optical immunosasensors)…………...25 1-4 碳六十化學…………………………………………………...27 1-4-1 碳六十的發現…………………………………………..…….27 1-4-2 碳六十的基本性質……………………………………..…….28 1-4-3 碳六十的化學反應…………………………………………...32 1-5 光敏電阻……………………………………………………...33 1-6 壓電晶體……………………………………………………...36 1-6-1 壓電晶體之壓電性……………………………………….36 1-6-2 石英振盪器………………………………………………….36 1-6-3 石英微量天平(Quartz crystal microbalance, QCM)…….39 1-6-4 石英振盪器的線路…………………………………………….43 1-6-5 石英壓電晶體在分析化學領域上的應用…………………….45 1-7 訊號對雜訊的加強(signal-to-noise enhancement)………..50 1-7-1整體平均………………………………………………………….50 1-7-2箱型平均………………………………………………………….51 第二章 實驗部分………………………………………………………52 2-1藥品與儀器…………………………………………………………52 2-1-1 藥品…………………………………………………………...52 2-1-2 使用儀器………………………………………………...52 2-2石英晶體的處理……………………………………………………53 2-2-1石英電極的表面塗佈液(coating solution)………………..53 2-2-2表面塗佈法……………………………………………………..54 2-3 實驗系統…………………………………………………………..54 2-3-1 光學偵測系…………………………………………………...54 2-3-1-1光學偵測原理…………………………………………….54 2-3-1-2 自製光學偵測系統簡述…………………………………55 2-3-2 QCM系統…………………………..…………………………59 2-4 實驗裝置與實驗步驟…………………………………………..61 2-4-1 以光學方式偵測抗體抗原…………………………………….61 2-4-2 碳六十與酵素-嗎啡間作用力的研究…………………………62 2-4-3抗嗎啡抗體與酵素-嗎啡之間結合的研究…………………….63 第三章 實驗結果與討論………………………………………………64 3-1光學偵測系統偵測嗎啡研究……………………………………..64 3-1-1抗嗎啡抗體與酵素-嗎啡溶液間作用………………………....64 3-1-2嗎啡-酵素及嗎啡抗體用量對電壓變化的影響………………70 3-1-3嗎啡濃度對於電壓改變的影響……….…….…………...……74 3-1-4光源對測量結果影響與光學感測器的再現性……..………...77 3-1-5體內常見干擾物對光學感測器的影響………….……………79 3-1-6 pH值對於偵測結果的影響…………………………………81 3-2利用石英壓電晶體偵測抗體………………………………………82 3-2-1碳六十與酵素-嗎啡間的作用力…………………………………82 3-2-2嗎啡抗體與嗎啡-酵素-之間的作用力……………….……….84 3-2-3碳六十/PVC塗佈量對嗎啡-酵素的效應…………………..……88 3-2-4嗎啡-酵素濃度對於C60/PVC塗佈壓電晶片頻率變化改變的影響………………………………………………………………………..89 3-2-5抗嗎啡抗體濃度對頻率改變所造成的影響………………….…90 3-2-6石英壓電晶體感測器的再現性………………………………….91 3-2-7血液中干擾物對壓電感測器的影響…………………………….92 第四章 結論……………………………………………………………94 參考文獻…………………………………………………………………a 附錄………………………………………………………………………e

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