血清中苯妥英治療藥物與壓阻微懸臂樑晶片偵測之研究

Abstract

苯妥英為近來使用最廣泛的抗癲癇藥物之一，其可藉由降低大腦電訊號的傳遞，抑制大腦不正常活動，進而達到避免癲癇發作的效果。苯妥英的治療劑量範圍很小而難以控制，若治療劑量過高，可能會導致藥物中毒；若劑量太低，則無法產生藥效。因此，須將藥物控制在適當的濃度方能充分發揮治療效果。治療藥物監測是一個觀察血液中苯妥英濃度的重要方法。隨著微機電系統(Micro-electromechanical System，MEMS）技術的進步，可攜帶、靈敏且可靠的壓阻式微懸臂樑生物感測器已被發展出來，其整合微流道系統後，可作為一免螢光標定之檢測平台。 為了能夠在胎牛血清（Fetal Bovine Serum，FBS）這種複雜的體液環境中測量苯妥英的濃度，需先量測苯妥英抗體和藥物的等電點（Isoelectric point，pI），以確定使用的FBS 其pH值應調整的範圍，而根據測量結果選用pH值8.05的50%及100% FBS來檢測100μg/ml苯妥英。使用壓阻式懸臂樑生物感測器時須先固定自組裝單層膜（Self-assembly Monolayer，SAM）和苯妥英抗體，最後加入藥物進行量測。在本研究中，首先測量在去離子（Deionized Water）水環境中，壓阻式微懸臂樑對10μg/ml苯妥英之反應性，得到解離常數（Dissociation Constant，KD）為58 μg/ml。接著測量100μg/ml苯妥英在pH 8.5的50%和100% FBS之反應，結果分別為0.36和0.24 N/m。此外，壓阻懸臂樑生物感測器的實驗結果，與傳統之螢光偏振免疫法（Fluorescence Polarization Immunoassay，FPIA）具有很高的一致性。

Phenytoin is one of the most widely used antiepileptic drugs recently. Its function is to suppress the abnormal brain activity seen in seizure by diminishing electrical conductance. However, its therapeutic dosage range in a circulatory system is very narrow and hard to be controlled. Otherwise, adverse effects may occur if the treatment dosage is too high, or an ineffective treatment may happen if the dosage is too low. So, only appropriate dosage must be given to reach a sufficient remedy. Therapeutic drug monitoring (TDM) is a crucial method to observe the concentration of phenytoin in blood vessel. With advances in micro-electromechanical system technology, a portable, sensitive and reliable piezoresistive-based microcantilever biosensor is fabricated and integrated with a microfluidic system for a label-free fluorescence detection. In order to measure phenytoin in fetal bovine serum (FBS) which is complex body fluid environment, the isoelectric points (pI) of phenytoin antibody and drug were measured to determine the window of pH value of FBS. As a result, the pH 8.05 of 50% and 100% FBS were selected to detect 100 μg/ml phenytoin. The piezoresistive microcantilever biosensor was utilized by first immobilizing a self-assembly monolayer (SAM) and followed by phenytoin antibody. Next, the biosensor was approved to detect phenytoin drug with 10 μg/ml limit of detection under deionized water environment. The dissociation constant (KD) was 58 μg/ml and the detection linear range was between 10 – 75 μg/ml. The concentration sensitivity was 2.94 х 10-6 (μg/ml)-1. Then, 100μg/ml phenytoin drug was also detected in the 50% and 100% FBS with adjusted pH8.05. Besides, the piezoresistive microcantilever biosensor showed an outstanding experimental agreement with a fluorescence polarization immunoassay (FPIA).