陳頌方Chen, Sung-Fang簡子傑Chien, Zu-Chieh2022-06-082026-07-202022-06-082021https://etds.lib.ntnu.edu.tw/thesis/detail/ab6075cbb0e7431a58b764089d286316/http://rportal.lib.ntnu.edu.tw/handle/20.500.12235/117235人類血清樣品包含多種蛋白質和核酸，其中許多生物標誌物是檢測疾病時非常重要的指標，然而此類低豐度生物標誌物的分析會受到高豐量人類血清白蛋白 (HSA)的干擾。本篇研究目的為開發一種有效分離HSA的材料，能夠重複利用、同時短時間內完成人類血清之前處理，最終使分析檢體中HSA的豐度降低。本篇研究利用表位印記方法 (epitope approach)合成分子印記聚合物 (Molecularly Imprinted Polymers, MIPs)，相較於過去許多文獻使用整個蛋白質，本篇以同於人類血清白蛋白N端之胜肽鏈作為模板分子，再以Methacrylic acid (MAA)為官能基單體、Ethylene glycol dimethacrylate (EGDMA)為交聯劑、2,2′-Azobis(2-methylpropionitrile) (AIBN) 為起始劑，並使用MeOH作為致孔溶劑合成出分子印記聚合物。且利用傅立葉紅外光譜儀 (FT-IR)、掃描式電子顯微鏡 (SEM) 對MIPs進行結構分析與外型觀測，優化模板分子、單體與交聯劑的比例，結果顯示當使用MKWVTFISLLFL作為模板分子，且模板/單體/交聯劑比例為1:21:85時，MIPs會具有最高的吸附能力 (1.026 µmol/g)、優異的選擇性 (Imprinting factor=2.93)，並且能快速地捕捉HSA (30分鐘)。最後成功證明此材料MIPs能從人類血清樣品中選擇性去除HSA，為人類血清前處理提供了一種快速且可重複使用的方法。Human serum samples contain diverse proteins and circulating nucleic acids, and some of them play important roles in disease prediction. However, the analysis of such low-abundance biomarkers is often hindered by abundant human serum albumin (HSA). The study developed a material which can be reused and conducted rapidly for human serum albumin depletion. In this study, a novel molecularly imprinted polymer was successfully prepared by epitope approach technique. N-terminus peptide sequence was selected as a test template molecule; Methacrylic acid (MAA), Ethylene glycol dimethyl acrylate, 2,2′-Azobis (2-methylpropionitrile) were served as the functional monomer, crosslinker, and initiator, respectively. Methanol was served as the porogen for the preparation process. Fourier transform infrared spectroscopy and scanning electron microscopy were applied to characterize the final product of molecularly imprinted polymers. The results indicated that excellent selectivity for HSA was obtained using MKWVTFISLLFL as template. The template/functional monomer/crosslinking agent at a ratio of 1:21:85 possess high adsorption capacity (1.026 µmol/g), superior selectivity (Imprinting factor = 2.93), and rapid capturing kinetics for HSA (30 min). Finally, it was successfully proved that this MIP can selectively remove HSA from human serum samples, providing a fast and reusable method for the pretreatment of human serum.人類血清白蛋白分子印記聚合物表位印記技術Human serum albuminMolecularly imprinted polymersEpitope approach學術論文