非酵素式葡萄糖酸鹼溶氧感測器之研發於幹細胞培養之監測應用

Abstract

幹細胞培養已有數十年歷史，這種細胞除了具有無限複製的能力，更能分化成各種不同功能的細胞。因此，人們對於其未來在醫學上的應用寄予厚望。然而，要維持幹細胞不分化或是分化形成特定細胞受環境影響甚鉅，這些影響包含物理性的刺激及化學性的刺激。如何利用這些刺激去分化出大量且純度高的細胞是研究人員長久以來極感興趣的議題也是目前所遇到的困難點。因此，若能發展出一套穩定的監測系統能長時間持續監測整個分化過程中培養基裡各種分子的濃度變化，不但能提供更多線索幫助研究者在各種機轉上的研究，更能幫助研究者更穩定控制細胞培養的環境克服現有的困難。 本論文的目的在於研究適合用於幹細胞培養長時間持續監測的葡萄糖感測器、酸鹼感測器及溶氧量感測器。由於幹細胞分化的過程所需的時間比一般細胞培養的時間來得長，因此在本研究中的感測器均採用非酵素式的電化學感測器。其中葡萄糖感測器是利用具奈米結構的鉑銥為感測材料，酸鹼感測器是利用氧化銥為感測材料，溶氧感測器是利用具奈米結構的鉑為感測材料。在感測靈敏度上，葡萄糖感測器為9.7 uA/cm2-mM，酸鹼感測器為68.5 mV/pH，溶氧感測器為28.7 uA/cm2-1% oxygen tension。並且，這些感測器的線性感測區間符合作為細胞培養液感測的需求。針對這三種感測器的感測原理、製作過程及各項功能測試將在本文中的各章節更詳細的敘述。

Stem cell research is one of the most fascinating areas of contemporary biology as stem cells are capable of dividing and self-renewing for long periods and have potential to give rise to any type of somatic cells. These unique properties allow us to generate specialized adult human cells for drug screening, as well as for replacing damaged tissue in cell therapy. However, either to produce a great deal of stem cells undifferentiated or to direct their differentiation into a specific cell lineage with high efficiency is still a challenge. Stem cell differentiation is influenced by a complex system, including ECM composition, medium additives (proteins or small molecules) and mechanical factors in the environment, etc. During the differentiation process, cell morphology is monitored on daily basis and gene/protein expression is checked at selected time points to certify the status of the progeny. It is useful if scientists can have thorough learning on the cell culture environment of differentiation process and, further, have control to make it favorable for the desired induction lineage. In this dissertation, the purpose is to develop three sensors, glucose, pH and oxygen sensors, for continuously monitoring the changes in the culture medium during stem cell differentiation process. The glucose, pH, and oxygen sensors have been made and are all nonenzymatic electrochemical sensors. The sensing materials of the glucose, pH and oxygen sensors are nanostructured Pt-Ir, IrO2-x, and nanostructured Pt, respectively. The sensitivities of the glucose, pH and oxygen sensors are 9.7 uA/cm2-mM, 68.5 mV/pH and 28.7 uA/cm2-1% oxygen tension, respectively. This dissertation explains the mechanism, the fabrication method and the characteristics of the three sensors’ functionality in each chapter.