拋棄式定量化學發光側向流動免疫蛋白檢測之開發

Abstract

對已開發或開發中國家而言，阿茲海默的患病率都在迅速增加，並且針對病情的藥物其療效幾乎都不甚理想，故目前阿茲海默等疾病治療極為困難。有文獻提到阿茲海默症這類腦疾病之發病率與體內DAO (D-Amino Acid Oxidase) 蛋白濃度呈正相關，因此許多學者投入開發DAO蛋白抑制劑。為能決定個人抑制劑需下的DAO劑量，故開發出定量DAO蛋白濃度的平台就顯得急迫也具重要性。 本論文使用側向流動之免疫測定 (LFIA) 在血漿中DAO蛋白濃度，並利用化學發光進行呈色。本設計使用酵素結合免疫吸附分析中 (ELISA) 的三明治法或是競爭法免疫測定，根據抗體或是抗原情況做調整。為了符合臨床需求，此系統提供0ng/mL-600ng/mL的濃度範圍之檢測，涵蓋了病人與健康人的DAO濃度。檢測方法使用化學冷光法，並使用特定應用程式定出正確濃度 (比色法) ，讓紙張型傳感器也能擁有定量的功能。試驗結果確認其平台能分辨出血漿中含有不同濃度的DAO蛋白，並趨勢與標準曲線相符，而且相對於標準ELISA流程，減少了一半以上檢測時間與人為操作的不便，故未來希望能在臨床檢測上使用。

In developing countries and developed counties, the prevalence of Alzheimer‘s disease is increasing rapidly. The efficacy of drugs for the disease cannot cure or halt disease progression. Literature suggest that the incidence of brain-related diseases such as Alzheimer's disease is positively correlated with the concentration of D-Amino Acid Oxidase (DAO) in the body promoting development of a series of DAO protein inhibitors. Consequently, there is an urgent need to develop a platform to determine the concentration of DAO protein in order to assess efficacy of DAO inhibitors. In this thesis, lateral flow immunoassay (LFIA) was used to detect the concentration of DAO protein in human plasma. The device utilizes enzyme-linked immunosorbent assay (ELISA) sandwich or competitive immunoassays based on antibody or antigen conditions. To meet clinical needs, the system provides a range of concentrations from 0 ng/mL to 600 ng/mL, covering DAO concentrations in patients and healthy individuals. Chemiluminescence was used for detection to quantify the protein concentration. Results show the trend is comparable with the standard ELISA process. The detection time is reduced by more than half with substantially reduced operator intervention. It is hoped that the device can benefit patients in clinical setting.