臺灣博碩士論文加值系統

近年來我國邁入高齡化社會，國人對衛生保健與食品安全更加重視，而生物感測器的發展日漸蓬勃，應用亦日漸廣闊，其快速檢測、高靈敏度、可攜式且操作簡便等特點吸引許多研究人員將其應用於臨床檢驗與居家照護等領域。肌酸酐 (Creatinine) 為肌酸 (creatine) 與磷酸肌酸 (creatine phosphate) 之代謝廢物，是人類血液與尿液中用來評估腎臟功能的重要指標。
本研究以肌酸酐作為模版分子，NVP (N-vinylpyrrolidone) 以及NVI (1-vinylimidazole) 作為功能性單體進行共聚合形成高分子薄膜，利用萃洗過程將肌酸酐從高分子中洗脫，產生與肌酸酐具有特異性吸附效果之模印高分子 (molecularly imprinted polymer, MIP) 薄膜電極。
本研究以 FT-IR 之官能基特徵峰存在與否驗證肌酸酐成功地從高分子膜洗脫出；並以 SEM 分析各階段修飾電極的表面型態；以循環伏安法 (cyclic voltammetry) 探討各階段修飾電極電化學活性面積的變化，並以 EIS 數據模擬出等效電路；透過感測環境與電極修飾條件的最適化，並利用交流阻抗頻譜對肌酸酐作定量檢測，其濃度對應阻抗變化率在10~120 mg/dL呈高度線性，即使在標準尿中於10~300 mg/dL的範圍亦呈高度線性。本研究開發之肌酸酐感測器對人類尿液檢體進行感測與比對，由檢體測量值可確認此生物感測器於臨床檢驗的可行性。

SUMMARY
Creatinine, the final product of the metabolism of creatine in mammals that removed from the body by kidneys. It is a reliable indicator for kidney’s function in human’s blood and urine. Biosensors are already well established in modern analytical chemistry. They have become important tools for clinical diagnostics, environmental analysis, production monitoring, drug detection or screening. Synthetic biomimetic receptors like molecularly imprinted polymers (MIPs) have shown to be a potential alternative to biomolecules as recognition element for biosensing.
In this work, sensors based on poly(N-vinylpyrrolidone-co-1-vinylimidazole) for the selective determination of creatinine are fabricated by heated polymerization, We not only discuss the conclusion of creatinine in MIP by FTIR characterization and morphology, but also optimize creatinine calibration and detection by electrochemical impedance spectroscopy (EIS). Finally, the fabricated creatinine sensor was applied to the detection of urine specimens for feasibility test on clinical application.

Keywords: creatinine, molecularly imprinted polymers, electrochemical sensor, electrochemical impedance spectroscopy, urine specimens

Introduction
Creatinine, is the dehydrogenated form of creatine in ATP metabolism. The normal range of serum creatinine is 0.5~1.5 mg/dL; while in urine, the normal range is 39~259 mg/dL for males and 28~217 mg/dL for females, respectively. The value varies with human’s age, lifestyle and muscle mass. The concentration of creatinine is less affected by the variation of dietary, therefore creatinine in blood and urine is a fairly reliable indicator for clinical evaluation of renal function.
Clinical analysis of creatinine is routinely carried out using colorimetric Jaffé reaction and enzymatical reaction. However, Jaffé reaction suffers from the poor selectivity toward numerous metabolites containing carbonyl group found, such as proteins, glucose, bilirubin, and ascorbic acid. Although the introduction of enzymes may improve specificity, enzymes always suffer from instability, high cost and complex procedures for the immobilization.
In recent years, molecularly imprinted polymers (MIPs) have been used to develop the new generation of biological/chemical sensors due to their superiorities such as high adsorption capacity, high specific recognition of selected analyte, low cost and excellent reusability.
In this work, N-vinylpyrrolidone and 1-vinylimidazole as functional monomers were copolymerized in the presence of creatinine by heat polymerization. Based on the formation of hydrogen bonds of these functional monomers and creatinine, the fabricated electrode can be introduced to improve the specific binding affinity of creatinine. By the linearity between impedance change and creatinine concentration, we can apply in urine specimen detection.
MATERIALS AND METHODS
The gold working electrode was prepared by a sputter coater (sputter coater 108 auto, Cressington). Calculation of the amount of creatinine removed is measured by HPLC (SPD-10A; SIL-9A; C-R6A). Incorporation of creatinine in MIP matrix before and after extraction was compared by FTIR. Morphology of the modified electrodes were observed by SEM images. All electrochemical experiments were performed by using (Electrochemical analyzer) AUT85707 and Zive SP1 controlled and acquired by electrochemical signals of the sample solution. Clinical urine specimens were collected from NCKU hospital for detection test by the MIP modified electrodes.
RESULTS AND DISCUSSION
The proportion of creatinine removed from the MIP film was measured by an HPLC and calculated as 80.2%, which showed good imprinting as well as removal efficiency. FTIR analysis for the MIP films before and after template removal, the peaks at 3,250 cm1 which is amide group (NH stretching) disappeared. Meanwhile, the CN group at 1,108 cm1 decreased, which showed creatinine was removed from the polymer.
From the EIS (electrochemical impedance spectroscopy) simulation of modified electrode, the mechanism of creatinine detection can be confirmed. Because of formation of cavities inside the polymeric film, fascilitating the electron transfer between background electrolyte and working electrode, then with the injection of creatinine solution, creatinine can be rebinded to recognition sites in the cavities, blocked the electron transfer and resulted in the increase of impedance of whole electrode.
The as-prepared MIP electrode was examined by electrochemical detection. The gold electrode was modified by allyl mercaptan and imprinted poly(N-vinylpyrrolidone-co-1-vinylimidazole) was synthesized onto the electrode with the optimal ratio (NVP:NVI= 4:1), The sensing background was optimized at pH 7.4 to get the best calibration curve. Linear calibration curve with a slope of 0.024 ·mg1dL was thus obtained with R2= 0.9971. By comparison between MIP and NIP (Non-molecularly imprinted polymer), the imprinting factor was 12, the modified electrode showed the specific binding towards creatinine molecules. Real urine sample test was accomplished by the modified electrode and HPLC, respectively. Comparison to these methods, the result is good enough for clinical detection and it preliminary feasibility in point of care (POC) detection.
The sensing performance of the MIP electrode in this work such as reusability and storage stability was also investigated, discussing the reusability and storage stability of this MIP electrode. After reused for 50 times, the MIP electrode still remains remarkable stability compared to the same electrode prepared in the very beginning.
Conclusion
The extraction process to remove creatinine from the polymer matrix is confirmed from HPLC measurement and FTIR analysis. Both the preparation of MIP electrode and detection of creatinine were optimized. The as-prepared MIP electrode can detect urine creatinine within the range of 10~120 mg/dL versus the electrochemical impedance change with a sensitivity of 0.024 ·mg1dL and a LOD of 5.68 mg/dL. Additionally, the MIP electrode thus prepared can be applied to the determination of creatinine concentration in urine specimens.

摘要 II
目錄 III
表目錄 IX
圖目錄 X
第一章 緒論 1
1-1 前言 1
1-1-1 生物感測器 (Biosensor) 1
1-1-2 模印高分子 (Molecularly imprinted polymer, MIP) 2
1-2 研究動機 2
第二章 文獻回顧 3
2-1 生物感測器 3
2-1-1 生物識別元件 (Bioreceptor) 3
2-1-2 生物感測器種類 5
2-2 模印高分子 (Molecularly imprinted polymer, MIP) 10
2-2-1 模印高分子之組成 10
2-2-2 分子模印方式 12
2-2-3 模印高分子製備方法 13
2-2-4 模印高分子之應用 14
2-3 肌酸酐 (Creatinine) 15
2-3-1 簡介 15
2-3-2 肌酸酐與腎臟疾病 15
2-3-3 臨床評估 16
2-3-4 肌酸酐檢測法 16
2-4 電化學方法 17
2-4-1 電化學感測反應 17
2-4-2 伏安法 18
2-4-3 計時電流法 (Chronoamperometry) 19
2-4-4 開環電位法 (Open-circuit potential，OCP) 19
2-4-5 電化學阻抗頻譜法 (Electrochemical impedance spectroscopy，EIS) 20
2-5 肌酸酐模印高分子之感測 22
第三章 實驗方法、材料與儀器 23
3-1 製備肌酸酐模版高分子電極 23
3-1-1 製備硫醇修飾之工作電極 23
3-1-2 製備模印高分子預聚合液 23
3-1-3 製備模印高分子薄膜電極 23
3-1-4 模印高分子薄膜電極之萃洗 24
3-2 萃洗MIP電極條件之最佳化 25
3-2-1以 HPLC 製備肌酸酐標準尿溶液之檢量線 25
3-2-2 萃洗液之選擇 25
3-3 電極之電化學特性分析 25
3-3-1電化學活性面積分析 25
3-3-2 電化學阻抗頻譜 (Electrochemical impedance spectroscopy, EIS) 分析 26
3-4 MIP電極之電化學系統分析 26
3-4-1 以交流阻抗分析法進行肌酸酐檢測 26
3-4-2 背景pH 27
3-5 MIP電極之修飾最佳化 27
3-5-1 共聚合單體比例之最佳化 27
3-6 以標準尿作為注射樣品背景溶液之感測 27
3-7 干擾物測試 27
3-8 相似物測試 27
3-9 臨床尿液檢體檢測 28
3-10 電極保存穩定性測試 28
3-11 電極重複使用性測試 28
3-12 傅立葉轉換紅外線光譜儀 (Fourier Transform Infrared Spectrometer, FT-IR) 28
3-13 掃描式電子顯微鏡 (Scanning Electron Microscope, SEM) 29
3-14 實驗藥品 30
3-15 實驗儀器 31
第四章 結果與討論 32
4-1 肌酸酐模印高分子之合成 32
4-1-1單體選擇 32
4-1-2 非模印高分子 (Non-molecularly imprinted polymer, NIP) 之合成 32
4-2 萃洗模印高分子電極 33
4-2-1 萃洗程序之最適化 33
4-2-2 以FT-IR測定萃洗前後模印高分子之差異 35
4-2-3 掃描式電子顯微鏡 (SEM) 對電極表面分析 36
4-3 模印高分子電極之電化學性質分析 37
4-3-1 預聚液導電性分析 37
4-3-2 電化學活性面積分析 38
4-3-3電極EIS與等效電路分析 41
4-4 肌酸酐之交流阻抗分析與感測 44
4-4-1 肌酸酐之電化學性質 44
4-4-2 肌酸酐感測機制 45
4-4-3 感測環境與設置條件 46
4-4-4 電極製備之條件確立 54
4-4-5 偵測極限與標準尿測試 58
4-5 選擇性探討 59
4-5-1 干擾物測試 59
4-5-2 相似物測試 62
4-6 臨床檢體檢測 63
4-7 電極穩定性測試 66
4-8 電極重複使用性測試 67
4-9 本研究與過去文獻比較 68
第五章 結論 70
參考文獻 71
表目錄
表 2-3-1 美國腎臟基金會 (K-DOQI) 對慢性腎衰竭之分期 [40] 16
表 2-5-1 肌酸酐感測之模印高分子電極相關文獻整理。 22
表 3-7-1 尿液成分之濃度。 27
表 4-2-1 萃洗時間與肌酸酐萃洗量之對應表。 34
表 4-2-2 攪拌速率與肌酸酐萃洗量之對應表。 35
表 4-2-3 肌酸酐官能基對應之 FT-IR 吸收波長。 36
表 4-2-4 萃洗後 MIP 官能基對應之 FT-IR 吸收波長。 36
表 4-3-1 各電極之RandlesSevcik 方程式、電化學活性面積與粗糙因子列表。 39
表 4-3-2 各階段電極模擬之等效電路元件參數。 44
表 4-4-1 不同交流頻率下肌酸酐感測結果比較。 46
表 4-4-2 不同攪拌速率之肌酸酐感測結果比較。 48
表 4-4-3 不同萃洗液之肌酸酐感測結果比較。 48
表 4-4-4 不同pH 值背景溶液之肌酸酐感測結果比較。 49
表 4-4-5 不同前置平衡時間之肌酸酐檢量線與線性關係比較。 50
表 4-4-6 單針與連續注入之比較。 52
表 4-4-7 比較單針與連續操作感測肌酸酐之結果。 53
表 4-4-8 不同硫醇修飾之MIP 電極對肌酸酐感測之結果。 55
表 4-4-9 不同單體比例聚合之MIP 電極對肌酸酐感測之結果。 57
表 4-4-10 不同肌酸酐量聚合之MIP 電極對肌酸酐感測之結果。 58
表 4-5-1 單成分單針選擇性測試之比較。 60
表 4-5-2 混合成分單針選擇性測試之比較。 61
表 4-5-3 單成分單針選擇性測試之比較。 63
表 4-5-4 混合成分單針選擇性測試之比較。 63
表 4-6-1 尿液臨床檢體之肌酸酐濃度分區比較。 66
表 4-7-1 電極穩定性測試。 66 
圖目錄
圖1-1-1臨床可監測的疾病診斷方法之示意圖。[1] 1
圖1-1-2生物感測器之示意圖。 1
圖1-1-3 模印高分子流程之示意圖。 2
圖2-1-1以肌酸酐酶分解產生肌酐來間接檢測肌酸酐之示意圖。[3] 3
圖2-1-2以一級與二級抗體組成之感測器示意圖。[5] 4
圖2-1-3以DNA探針組成之感測器與其核酸序列示意圖。[7] 4
圖2-1-4 化學接受器型 (親離子團) 與肌酸酐結合之示意圖。[9] 5
圖2-1-5 以SELEX 篩選aptamer之流程圖。[10] 5
圖2-1-6 辨識元件固定化之示意圖。[11] (a) 物理吸附法；(b) 包埋法；(c) 共價接合法；(d) 親和作用法 6
圖2-1-7 包埋法。[12] (a) 以PPy作為包埋法之高分子示意圖；共價接合法。[13] (b) 以EDC/NHS固定化示意圖 7
圖2-1-8 親合作用法。[14] (a) Avidin-biotin親和作用法示意圖；分子模印。[16] (b) 模印高分子示意圖 7
圖2-1-9 場效電晶體示意圖。[18] (a) ISFET與模印高分子結合圖；[19] (b) 以肌酸酐做模版製備模印高分子固定於ISFET之示意圖 8
圖2-1-11 表面電漿共振示意圖。 [22] 8
圖2-1-12 局部表面電漿共振所引發之集體震盪現象 [23,24]。 9
圖2-1-13 拉曼能階變化示意圖 [25]。 9
圖2-1-14 全像攝影感測器。[26] (a) 全像攝影感測示意圖；[27] (b) 葡萄糖凝膠於全像攝影感測結果 10
圖2-1-15 智慧型手機感測。[28] (a)以智慧型手機進行感測之示意圖；[29] (b) 將模印高分子修飾之感測晶片應用於智慧型手機感測 10
圖2-2-1非共價性與共價性模印高分子之示意圖。[32] 11
圖 2-3-1 肌酸與肌酸酐之代謝途徑。 15
圖 2-3-2 Jaffé method 之反應式。 17
圖 2-4-1 電化學感測反應。[51] ( a) 電雙層示意圖；(b) 電化學反應路徑 18
圖 2-4-2 標準循環伏安法示意圖。(a) 施加電壓之波形變化圖；(b) 循環伏安圖 18
圖 2-4-3 脈衝電壓變化示意圖。(a) 微分脈衝伏安法；(b) 方波伏安法 19
圖 2-4-4 電化學分析之阻抗分析。[11] (a) 等效電路圖；[54] (b) Nyquist 圖 21
圖 2-4-5 肌酸酐模印高分子之交流阻抗式感測。[59] (a) 不同濃度之肌酸酐Nyquist 圖；(b) 肌酸酐定量之檢量線 22
圖 3-1-1 本研究肌酸酐模印高分子生物感測器之進行架構圖。 23
圖 3-1-2 肌酸酐MIP製備之示意圖。 24

圖 3-2-1 肌酸酐標準尿溶液之HPLC檢量線。 25
圖 3-2-2 以MIP電極進行肌酸酐交流感測系統示意圖。 26
圖 3-8-1 肌酸酐相似物之化學結構。(a) NHS (N-hydroxysuccinimide)；(b) 2-pyrrolidinone 28
圖 4-1-1 MIP 與 NIP 電極吸附機制差異圖。 32
圖 4-2-1 萃洗時間最適化條件之選擇。(a) 15分鐘；(b) 30 分鐘；(c) 45 分鐘；(d) 60 分鐘 33
圖 4-2-2 攪拌速率最適化條件之選擇。(a) 0 rpm；(b) 60 rpm；(c) 100 rpm；(d) 120 rpm 34
圖 4-2-3 肌酸酐與萃洗前後模印高分子之FT-IR 圖譜差異。(a) 肌酸酐；(b) 萃洗前之 MIP 電極；(c) 萃洗後之 MIP 電極；(d) 肌酸酐與萃洗前後 MIP 電極 36
圖 4-2-4 金電極之SEM 影像。(a) 俯視圖 (×1.0 k)；(b) 側視圖 (×400) 37
圖 4-2-5 萃洗前 MIP 電極之 SEM 影像。(a) 俯視圖 (×1.0 k)；(b) 側視圖 (×400) 37
圖 4-2-6 萃洗後 MIP 電極之 SEM 影像。(a) 俯視圖 (×1.0 k)；(b) 側視圖 (×2.0 k) 37
圖 4-3-1 預聚合液與其溶劑之導電分析。 38
圖 4-3-2 各電極之電化學活性面積圖。 39
圖 4-3-3 不同階段之修飾電極於不同掃描速率下之CV圖。(a) 裸金電極；(b) 萃洗前之MIP電極；(c) 萃洗後之MIP電極，與氧化還原電流峰值之線性關係；(d) 裸金電極；(e) 萃洗前之MIP電極；(f) 萃洗後之MIP電極。 40
圖 4-3-4 各修飾電極之EIS 分析。(a) Nyquist 圖；(b) Bode 圖之相位角對頻率 41
圖 4-3-5 各階段修飾電極之等效電路模擬 (左) Nyquist 圖；(右) Bode 圖之相位角對頻率。(a), (c) 裸金電極；(b), (d) MIP 電極。 42
圖 4-3-6 各階段電極之等效電路模擬與電極結構示意圖。(a) 裸金電極；(b) MIP 電極 43
圖 4-4-1 MIP電極加入肌酸酐之赤血鹽/黃血鹽水溶液CV圖。 44
圖 4-4-2 不同交流頻率下對肌酸酐感測之阻抗變化率。(a) 感測圖；(b) 阻抗變化率比較圖 46
圖 4-4-3 不同攪拌速率下對肌酸酐感測之阻抗對時間變化。(a) 0 rpm；(b) 60 rpm；(c) 100 rpm；(d) 120 rpm 47
圖 4-4-4 不同萃洗液對肌酸酐感測之影響。(a) 去離子水；(b) NaOH；(c) HCl 48
圖 4-4-5 不同萃洗液對肌酸酐之萃洗量。(a) 去離子水；(b) NaOH；(c) HCl 49
圖 4-4-6 不同pH值背景溶液下對肌酸酐感測之阻抗對時間變化。(a) pH= 7.0；(b) pH= 7.4；(c) pH= 8.0；(d) pH= 9.0 50
圖 4-4-7 不同前置平衡時間下對肌酸酐感測之阻抗變化率對時間變化。(a) 1,000 s；(b) 2,000 s；(c) 3,000 s，與肌酸酐濃度對阻抗變化率繪製之檢量線。(d) 1,000 s；(e) 2,000 s；(f) 3,000 s 51
圖 4-4-8 以一階系統響應模擬 MIP 修飾電極對肌酸酐感測之阻抗變化時間圖。 52
圖 4-4-9 以單針或連續注入之肌酸酐溶液感測之阻抗變化率對時間變化。(a) 單針注入，MIP 電極；(b) 單針注入，NIP 電極；(c) 連續注入，MIP 電極；(d) 連續注入， NIP 電極，與肌酸酐濃度對阻抗變化率繪製之檢量線。(e) 單針注入；(f) 連續注入 54
圖 4-4-10 不同硫醇修飾製作之 MIP電極對肌酸酐感測之阻抗對時間變化。(a) 2-mercaptoethanol；(b) allyl mercaptan，與其肌酸酐萃洗量。(c) 2-mercaptoethanol；(d) allyl mercaptan 55
圖 4-4-11 NVP與NVI共聚合之反應示意圖。[64] 56
圖 4-4-12 不同單體比例之 MIP 電極對肌酸酐感測之阻抗變化率對時間變化。NVP：NVI= (a) 1：0；(b) 4：1；(c) 1：1；(d) 1：4；(e) 0：1 57
圖 4-4-13不同肌酸酐量聚合之 MIP 電極對肌酸酐感測之阻抗變化率對時間變化。 (a) 10 mg；(b) 20 mg；(c) 30 mg 58
圖 4-4-14 標準尿測試。(a) 阻抗變化率對時間作圖；(b) 肌酸酐檢量線 59
圖 4-5-1 單成分單針選擇性測試。 60
圖 4-5-2 肌酸酐/干擾物混合成分單針選擇性測試。 61
圖 4-5-3 肌酸酐/干擾物混合成分多針選擇性測試。第一針與最後一針皆為肌酸酐：(a) 25、100 mg/dL；(b) 25、25 mg/dL；(c) 100、25 mg/dL，與 (d) 隨機注入肌酸酐或干擾物 62
圖 4-5-4 相似物測試。(a) 單成分相似物測試；(b) 肌酸酐/相似物測試 63
圖 4-6-1 全檢體統計圖。 64
圖 4-6-2 以 MIP 電極之臨床尿液檢測結果。(a) 與謝淑珠老師實驗室提供之數據(kinetic Jaffé method) 比較；(b) 與 HPLC 比較 64
圖 4-6-3將臨床檢體肌酸酐濃度作分區定量結果。(a) 0~50 mg/dL；(b) 50~100 mg/dL；(c) 100~150 mg/dL；(d) 150~200 mg/dL；(e) above 200 mg/dL 65
圖 4-7-1 電極穩定性測試。 66
圖 4-8-1 電極重複使用性測試。 67
圖 4-8-2 電極重複使用性測試與對應天數。 67

1.S Wang, T Chinnasamy, M Lifson, F Inci, U Demirci. Flexible substrate-based devices for point-of-care diagnostics. Trends in Biotechnology, 34, 909921, 2016.
2.J Wang. Glucose biosensors: 40 years of advances and challenges. Electroanalysis, 13, 983‒988, 2001.
3.FH Cincotto, EL Fava, FC Moraes, O Fatibello-Filho, RC Faria. A new disposable microfluidic electrochemical paper-based device for the simultaneous determination of clinical biomarkers. Talanta, 195, 6268, 2019.
4.MU Ahmed, MM Hossain, M Safavieh, YL Wong, IA Rahman, M Zourob, E Tamiya. Toward the development of smart and low cost point-of-care biosensors based on screen printed electrodes. Critical Reviews in Biotechnology, 36, 495‒505, 2016.
5.Z Zhou, D Du, J Wang, JN Smith, C Timchalk, Y Li, Y Lin. Quantum dot-based immunochromatographic fluorescent biosensor for biomonitoring trichloropyridinol, a biomarker of exposure to chlorpyrifos. Analytical Chemistry, 82, 5125‒5133, 2010.
6.教育部顧問室「生物技術科技教育改進計畫」分子檢驗。2003
7.S Lee, H Jo, J Her, H Lee, C Ban. Ultrasensitive electrochemical detection of engrailed-2 based on homeodomain-specific DNA probe recognition for the diagnosis of prostate cancer. Biosensors and Bioelectronics, 66, 32‒38, 2015.
8.M Subat, AS Borovik, B König. Synthetic creatinine receptor:  imprinting of a Lewis acidic zinc(II)cyclen binding site to shape its molecular recognition selectivity. Journal of American Chemical Society, 126, 3185‒3190, 2004.
9.T Guinovart, DH Alonso, L Adriaenssens, P Blondeau, FX Rius, P Ballester, FJ Andrade. Characterization of a new ionophore-based ion-selective electrode for the potentiometric determination of creatinine in urine. Biosensors and Bioelectronics, 87, 587‒592, 2017.
10.S Song, L Wang, L Wang, J Li, J Chao, C Fan. Aptamer-based biosensors. Trac-Trends in Analytical Chemistry, 27, 108‒117, 2008.
11.N Aydemir, J Malmstrom, J Travas-Sejdic. Conducting polymer based electrochemical biosensors. Physical Chemistry Chemical Physics, 18, 8264‒8277, 2016.
12.F Wei, S Cheng, Y Korin, EF Reed, D Gjertson, C Ho, HA Gritsch, J Veale. Serum creatinine detection by a conducting-polymer-based electrochemical sensor to identify allograft dysfunction. Analytical Chemistry, 84, 7933‒7937, 2012.
13.A Diouf, S Motia, NE Hassani, NE Bari, B Bouchikhi. Development and characterization of an electrochemical biosensor for creatinine detection in human urine based on functional molecularly imprinted polymer. Journal of Electroanalytical Chemistry, 788, 4453, 2017.
14.GM Holzinger, AL Gof, S Cosnier. Supramolecular immobilization of bio-entities for bioelectrochemical applications. New Journal of Chemistry, 38, 5173-5180, 2014.
15.J Ballesta-Claver, J Ametis-Cabello, J Morales-Sanfrutos, A Megía-Fernández, MC Valencia-Mirón, F Santoyo-González, LF Capitán-Vallvey. Electrochemiluminescent disposable cholesterol biosensor based on avidin–biotin assembling with the electroformed luminescent conducting polymer poly(luminol-biotinylated pyrrole). Analytica Chemica Acta, 91‒98, 2012.
16.A Zamora-Galvez, E Morales-Narvaez, CC Mayorga-Martinez, A Merkoci. Nanomaterials connected to antibodies and molecularly imprinted polymers as bio/receptors for bio/sensor applications. Applied Materials Today, 9, 387‒401, 2017.
17.M Dabrowski, M Cieplak, PS Sharma, P Borowicz, K Noworyta, W Lisowski, F D'Souza, A Kuhn. Hierarchical templating in deposition of semi-covalently imprinted inverse opal polythiophene film for femtomolar determination of human serum albumin. Biosensors and Bioelectronics, 94, 155‒161, 2017.
18.R Schirhagl. Bioapplications for Molecularly Imprinted Polymers. Analytical Chemistry, 86, 250‒261, 2014.
19.H Tsai, C Lin, Y Juang, I Wang, Y Lin, R Wang, H Lin. Multiple type biosensors fabricated using the CMOS BioMEMS platform. Sensors and Actuators B, 144, 407412, 2010.
20.M Syu, T Hsu, Z Lin. Synthesis of recognition matrix from 4-methylamino-N-allylnaphthal-imide with fluorescent effect for the imprinting of creatinine. Analytical Chemistry, 82, 8821‒8829, 2010.
21.J Lee. Bioluminescence, the Nature of the Light. University of Georgia Libraries, 2017.
22.Y Yanase, T Hiragun, K Ishii, T Kawaguchi, T Yanase, M Kawai, K Sakamoto, M Hide. Surface plasmon resonance for cell-based clinical diagnosis. Sensors, 14, 4948‒4959, 2014.
23.JC Wang, WH Lin, E Cao, XF Xu, WJ Liang, XF Zhang. Surface plasmon resonance sensors on raman and fluorescence spectroscopy. Sensors, 17, 2719, 2017.
24.S Unser, I Bruzas, J He, L Sagle. Localized surface plasmon resonance biosensing: current challenges and approaches. Sensors, 15, 15684‒15716, 2015.
25.https://en.wikipedia.org/wiki/Raman_spectroscopy.
26.R Ahmed, AK Yetisen, SH Yun, H Butt. Color-selective holographic retroreflector array for sensing applications. Light-Science ＆ Applications, 6, e16214, 2017.
27.M Elsherif, MU Hassan, AK Yetisen, H Butt. Glucose sensing with phenylboronic acid functionalized hydrogel-based optical diffusers. ACS Nano, 12, 2283‒2291, 2018.
28.D Zhang, Q Liu. Biosensors and bioelectronics on smartphone for portable biochemical detection. Biosensors and Bioelectronics, 75, 2016.
29.D Capoferri, R Álvarez-Diduk, M Del Carlo, D Compagnone, A Merkoçi. Electrochromic molecular imprinting sensor for visual and smartphone-based detections. Analytical Chemistry, 90, 5850‒5856, 2018.
30.S Zhang, R Geryak, J Geldmeier, S Kim, VV Tsukruk. Synthesis, assembly, and applications of hybrid nanostructures for biosensing. Chemical Reviews, 117, 12942‒13038, 2017.
31.R Gui, H Jin, H Guo, Z Wang. Recent advances and future prospects in molecularly imprinted polymers-based electrochemical biosensors. Biosensors and Bioelectronics, 100, 56‒70, 2018.
32.Y Fuchs, O Soppera, K Haupt. Photopolymerization and photostructuring of molecularly imprinted polymers for sensor applications—A review. Analytica Chemica Acta, 717, 7‒20, 2012.
33.G Wulff, J Haarer. The preparation of defined chiral cavities for the racemic resolution of free sugars. Markromol.Chem, 192, 1329‒1338, 1991.
34.H Rao, Z Lu, H Ge, X Liu, B Chen, P Zou, X Wang, H He, X Zeng, Y Wang. Electrochemical creatinine sensor based on a glassy carbon electrode modified with a molecularly imprinted polymer and a Ni@polyaniline nanocomposite. Microchim Acta, 184, 261‒269, 2017.
35.B Babamiri, A Salimi, R Hallaj, M Hasanzadeh. Nickel nanoclusters as a novel emitter for molecularly imprinted electrochemiluminescence based sensor toward nanomolar detection of creatinine. Biosensors and Bioelectronics, 107, 272279, 2018.
36.L N Gómez-Arribas, J L Urraca, E Benito-Peña, M C Moreno-Bondi. Tag-Specific Affinity Purification of Recombinant Proteins by Using Molecularly Imprinted Polymers. Analytical Chemistry, 91, 4100‒4106, 2019.
37.柯定賢。合成螢光單體以製備具螢光之模版高分子膜用於螢光式感測肌酸酐。國立成功大學化工系碩士學位論文，2009
38.X Zhang, F Zheng, L Ye, P Xiong, L Yan, W Yang, B Jiang. A one-pot sol-gel process to prepare a superhydrophobic and environment-resistant thin film from ORMOSIL nanoparticles. RSC Advances, 4, 9838‒9841, 2014.
39.J Coresh, BC Astor, T Greene, G Eknoyan, AS Levey. Prevalence of chronic kidney disease and decreased kidney function in the adult US population: Third National Health and Nutrition Examination Survey. American Journal of Kidney Diseases, 41, 1‒12, 2003.
40.吳明儒。評估腎臟功能的方法。腎臟語病析，19，45-49，2007
41.SR Dunn, GM Gabuzda, KR Superdock, RS Kolechi, RW Schaedler, ML Simenhoff. Induction of creatininase activity in chronic renal failure: Timing of creatinine degradation and effect of antibiotics. American Journal of Kidney Diseases, 29, 7277, 1997.
42.M Peake, M Whiting. Measurement of Serum Creatinine – Current Status and Future Goals. The clinical biochemist reviews, 4, 173‒184, 2006.
43.V Nguyen, C Wolff, J Seris, J Schwing. Immobilized Enzyme Electrode for Creatinine Determination in Serum. Analytical Chemistry, 63, 611614, 1991.
44.F Wei, S Cheng, Y Korin, E Reed, D Gjertson, C Ho, H Gritsch, J Veale. Serum Creatinine Detection by a Conducting Polymer Based Electrochemical Sensor to Identify Allograft Dysfunction. Analytical Chemistry, 84, 7933‒7937, 2012.
45.EP Randviir, CE Banks. Analytical methods for quantifying creatinine within biological media. Sensors and Actuators B: Chemical, 183, 239252, 2013.
46.EP Randviir, DK Kampouris, CE Banks. An improved electrochemical creatinine detection method via a Jaffé-based procedure. Analyst, 138, 6565‒6572, 2013.
47.AJ Killard, MR Smyth. Creatinine biosensors: principles and designs. Trends in Biotechnology, 18, 433‒437, 2000.
48.T Wen, W Zhu, C Xue, J Wu, Q Han, X Wang, X Zhou, H Jiang. Novel electrochemical sensing platform based on magnetic field-induced self-assembly of Fe3O4@Polyaniline nanoparticles for clinical detection of creatinine. Biosensors and Bioelectronics, 56, 180‒185, 2014.
49.Q Ang, F Chan, P Tan, S Low. Improving imprinted shape cavities of molecularly imprinted sol–gel host matrix with minimal relaxation for sensing of creatinine. Journal of Sol-Gel Science and Technology, 86, 226238,2018.
50.AJ Bard, LR Faulkner. Electrochemical methods: fundamentals and applications. 2001.
51.J Huang, Z Li, H Ge, JB Zhang. Analytical solution to the impedance of electrode/electrolyte interface in lithium-ion batteries. Journal of the electrochemical Society, 162, A7037A7048, 2015.
52.N Elgrishi, KJ Rountree, BD McCarthy, ES Rountree, TT Eisenhart, JL Dempsey. A practical beginner’s guide to cyclic voltammetry. Journal of Chemical Education, 95, 197‒206, 2018.
53.F Scholz, AM Bond, RG Compton, DA Fiedler, G Inzelt, H Kahlert. Electroanalytical methods: guide to experiments and applications. 2nd Ed., Springer, Inc., Berlin 2010.
54.D Pihíková, P Kasák, J Tkac. Glycoprofiling of cancer biomarkers: Label-free electrochemical lectin-based biosensors. Open Chemistry, 13, 636‒655, 2015.
55.E Barsoukov, JR Macdonald. Impedance spectroscopy theory, experiment, and applications. 3rd Ed., John Wailey＆ Sons, Inc., USA 2018.
56.JB Jorcin, ME Orazem, N Pébère, B Tribollet. CPE analysis by local electrochemical impedance spectroscopy. Electrochemica Acta, 51, 1473‒1479, 2006.
57.許庭榕。以分子模版技術用於螢光式與電化學阻抗式分析感測肌酸酐。國立成功大學化工系碩士學位論文，2010
58.黃宥霖。製備溶膠凝膠模版高分子薄膜電極用於肌酸酐濃度之交流阻抗式感測。國立成功大學化工系碩士學位論文，2015
59.KK Reddy, KV Gobi. Artificial molecular recognition material based biosensor for creatinine by electrochemical impedance analysis. Sensors and Actuators B: Chemical, 183, 356363, 2013.
60.B Khadro, C Sanglar, A Bonhomme, A Errachid, N Jaffrezic-Renault. Molecularly imprinted polymers (MIP) based electrochemical sensor for detection of urea and creatinine. Procedia Engineering, 5, 371374, 2010.
61.C Huang, T Tsai, J Thomas, M Lee, B Liu, H Lin. Urinalysis with molecularly imprinted poly(ethylene-co-vinyl alcohol) potentiostat sensors. Biosensors and Bioelectronics, 24, 26112617, 2009.
62.F Martinez-Pina, L Gargallo, D Radic. N-vinylimidazole-N-vinyl-2-pyrrolidone copolymers. Part I. reactivity ratios, solubility and viscosimetric study. Polymer International, 47, 340344, 1998.
63.S Ida, H Kitanaka, T Ishikawa, S Kanaoka, Y Hirokawa. Swelling properties of thermoresponsive/hydrophilic co-networks with functional crosslinked domain structures. Polymer Chemistry, 9, 17011709, 2018.
64.https://en.wikipedia.org/wiki/1-Vinylimidazole#cite_note-12
65.J Du, B Zhu, WR Leow, S Chen, TC Sum, X Peng, X Chen. Colorimetric detection of creatinine based on plasmonic nanoparticles via synergistic coordination chemistry. Small, 33, 4104‒4110, 2015.
66.CM Stumpe, H Grubmueller. Aqueous urea solutions: structure, energetics, and urea aggregation. Journal of Physical Chemistry B, 111, 62206228, 2007.