本研究利用半導體製程製備植入型微電極陣列探針，製程主要可分成三個階段，第一階段為微電極陣列金屬圖層之形成，利用微影製程技術及電子蒸鍍，將金屬鉑鍍在矽基材上。第二階段為微電極陣列探針表層的絕緣，此階段將探針沉積絕緣層除了電極端及封裝端以外，使用電漿輔助化學氣相沉積將其表面覆蓋介電層作絕緣，並將電極端與封裝端進行局部蝕刻以便後續研究。第三階段為微電極陣列探針邊界輪廓定義，利用微影製程定義出探針邊界輪廓，進行深蝕刻將探針從晶圓中取下。
取下微電極陣列探針後將進行其表面修飾，分別製備兩種感測器，第一種為以二茂鐵修飾電極製備穀氨酸感測器，將微電極陣列探針先沉積抗干擾層聚咯薄膜與全氟磺酸薄膜，在顯微鏡下以人工徒手塗佈將穀氨酸氧化酵素(利用戊二醛交聯劑和牛血清蛋白穩定劑與穀氨酸氧化酵素進行交聯)固定在電極表面，最後將與牛血清蛋白交聯二茂鐵(利用1-(3-二甲氨基丙基)-3-乙基碳二亞胺鹽酸鹽及N-羥基琥珀醯亞胺兩交聯劑混合二茂鐵和牛血清蛋白進行交聯)以物理吸附的方式固定於穀氨酸感測器上。第二種為以二茂鐵修飾電極製備過氧化氫感測器，將微電極陣列探針先沉積抗干擾層聚咯薄膜與全氟磺酸薄膜，最後將牛血清蛋白交聯二茂鐵以物理吸附方式固定於過氧化氫感測器上，其中，在製備過氧化氫感測器時毋須固定酵素，因為過氧化氫為電中性分子藉由擴散作用會穿越抗干擾層至白金電極表面，被氧化放出電子。
以二茂鐵所修飾的穀氨酸感測器，其線性範圍為20-539µM，響應時間約3秒，感測極限為1.35 µM (N = 30)，靈敏度為121 nA/µM·cm2(N = 3)，相較於未以二茂鐵所修飾的穀氨酸感測器，其靈敏度增加了1.51倍，儲存穩定性為123%(存放28天後其靈敏度與其初始靈敏度比值之百分比)。另外，以二茂鐵修飾電極製備過氧化氫感測器，其響應時間約2秒，靈敏度為269.0 nA/µM·cm2(N = 3)，相較於未以二茂鐵所修飾的過氧化氫感測器，其靈敏度增加了3.73倍。
本研究結合二茂鐵所修飾的穀氨酸感測器微電極陣列探針，以及未使用二茂鐵修飾的穀氨酸感測器微電極陣列探針，將此二種穀氨酸感測器探針同時植入白鼠大腦視丘中，分別刺激白鼠左肢、右肢，以偵測由痛覺刺激所產生的穀氨酸釋放，也探討以二茂鐵修飾之穀氨酸感測器用於活體實驗之效果，實驗結果發現動物體內神經傳導途徑為對側控制系統，另外有修飾二茂鐵的穀氨酸微電極陣列探針相較於未使用二茂鐵修飾的穀氨酸感測器微電極陣列探針，在動物實驗裡並沒有成功的放大穀氨酸訊號，其可能原因為血液中的血紅素與感測器上的二茂鐵結合造成感測器表面阻塞進而導致靈敏度下降。

In this research, the semiconductor manufacturing technology was used to fabricate implantable microelectrode array (MEA) probe. The manufacturing process can be divided into three parts. The first part is the formation of the metal layer on probes that defined electrode sites, connections, and bonding pads. The photolithography technology and metal deposition technology by electron beam evaporator were used to transfer the metal pattern on the silicon substrate. The second part is passivation process of the probe surface. The wafer surface were deposited with dielectric layers (SiO2 and Si3N4) by plasma enhanced chemical vapor deposition (PECVD). Then, the electrode sites and the bonding pads defined by the second photolithography process were etched to expose their metal surfaces. The third part is the definition of the probe outline. The third photolithography process was used to define the pattern of probe outline and then, the etching process was used to etch the outline to the bottom of the substrate in order to make the probes releasable from the wafer.
After the probes were released from the wafer, the electrode surface would be modified for fabricating glutamate sensors. In this research, there were two kinds of sensor would be prepared. The first one was the ferrocene modified microelectrode array glutamate sensor. First, permselective polymer layers (polypyrrole and Nafion®) for blocking interferents were deposited. Then, glutamate oxidase was immobilized on the electrode manually by crosslinking using the stabilizing reagent bovine serum albumin (BSA) and the crosslinker glutaraldehyde (GAH). Ferrocene conjugated BSA were prepared by using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS) crosslinkers mixed with ferrocene solution and BSA solution. Finally, the Fc-BSA was adsorbed on the sensor by immersing the sensor into the solution of ferrocene conjugate. The other sensor that we fabricated was the ferrocene modified microelectrode array hydrogen peroxide sensor. Similarly, the permselective polymer layers (polypyrrole and Nafion®) for blocking interferents were deposited. Finally, the Fc-BSA was adsorbed on the sensor by immersing the sensor into the solution of ferrocene conjugate. When fabricating the hydrogen peroxide sensor, no enzyme was required because of hydrogen peroxide was a electrical neutrality of molecule which could diffuse into electrode surface, It’s was be oxidased.
The glutamate sensors were modified by ferrocene have a larger linear detection range (20–539 μM), higher sensitivity 121 nA/µM·cm2(N = 3), it also have faster response time (<3 s), and lower detection limit (1.35 μM). The long-term stability of ferrocene modification glutamate sensor has been investigated by testing sensor sensitivities every two week for 28 days (sensors was stored in a desiccated box at -20°C). The ferrocene modification sensor retains 123 % of its original activity. Additionally, The hydrogen peroxide sensors were modified by ferrocene have higher sensitivity 269.0 nA/µM·cm2(N = 3), After modification the hydrogen peroxide sensor have 3.73 times sensitivity higher than the unmodified hydrogen peroxide sensor. It also have faster response time (<2 s).
In this research, the ferrocene modified microelectrode array glutamate sensors and the unmodified glutamate sensors were combined and implanted into the thalamus of rats together. The evoked glutamate release by noxious stimulation on the right leg and the left leg of rats were monitored. The performance of the ferrocene modified microelectrode array glutamate sensors applied in rats was evaluated. The results showed that the in vivo study confirmed that the ascending pathway of spinothalamic tract was contralateral. Additionally, the ferrocene modified microelectrode array glutamate sensors didn’t show the higher sensitivity in the animal test comparing with the unmodified glutamate sensors. The reasons were lacking of ferrocene in vivo and hemosiderin were attracted the iron in ferrocene. That were the reasons why ferrocene couldn’t work in animal test.

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