石英晶體微天平應用於人體免疫球蛋白檢測之實驗及模擬

Abstract

由於石英晶體微天平生物感測器的無須標定、無須破壞生物分子、高靈敏度和能夠即時檢測等優異特性，在近幾年來廣泛的應用於生物分子間反應行為的即時檢測上。關於石英晶體微天平在檢測生物分子行為的相關研究大多以實驗為主，利用數值模擬討論鍵結反應的相關研究卻略顯不足。本論文利用有限元素進行討論，並與實驗結果加以比對與驗證，相互映證彼此的正確性。 本研究利用石英晶體微天平進行人體免疫球蛋白(Human IgG1)和其相對應的抗原(Anti-Human IgG1)之親和解離行為的即時檢測及分析。再利用有限元素模擬其鍵結反應行為，並和實驗結果相互驗證。研究中發現溶液輸送流道中穩態層流的流場會影響流道內濃度的分布。當流道長度固定，待測物供應量越少，輸送流道出口處的濃度分佈會有明顯的衰減。研究中又發現由於質量傳輸限制導致親和力分析中未能滿足 的假設。上述的兩大原因都會使得在親和力分析中產生計算上的誤差，造成實驗與模擬結果會有明顯的差異。因此我們建立一套新的修正結合速率常數 和解離速率常數 計算方法，並利用修正過 、 與完整的模型進行反應曲線的模擬，其結果即獲得相當大的改善。模擬的反應曲線與實驗之反應曲線會有較佳的相似度，證明了修正的計算方法與模擬模型的正確性。最後利用此驗證過的模擬模型，對各項變數與生物鍵結反應之間的影響進行討論。

Quartz Crystal Microbalance (QCM) biosensor has several advantages in analyzing the interaction among biomolecules, such as label free, non-destructive, highly sensitive and capable of monitoring dynamic biomolecular interaction in real time. It has been widely used as the apparatus for the biomolecule detection in the last two decades. Most of the existing works are focused merely on experiments of using QCM. In this thesis, we perform not only on experiments but also the numerical simulation based on the FEM software, COMSOL Multiphysics, to study the behavior of the antibody-antigen interaction. In these immunoassay experiments, we use the biosensor (Quartz Crystal Microbalance, QCM) to detect the specific binding reaction of the Human IgG1-Anti-Human IgG1 protein pair in physiological environments. In addition to experiments, we use the finite element analysis software, COMSOL Multiphysics, to simulate the behavior of Human IgG1 and Anti-Human IgG1 interactions. During the simulation process, we discuss the unsteady convective diffusion in fully developed laminar flow in the transport tube. The transport tube is used for transporting the analyte solution into the microchannel. The analyte concentration in the tube is strongly affected by the flow field. With the supplement of the analyte solution decreasing, the analyte concentration will be apparently decayed when length of the tube is fixed. Furthermore, the assumption of [A]surface = [A]bulk in the basic kinetic analysis is not correct because of the effect by mass transport in the liquid phase. These above-mentioned reasons cause that the apparent association rate constant and the apparent dissociation rate constant obtained by the basic kinetic analysis are not the real constants of the specific binding reaction. The apparent and the apparent cause that results obtained from the simulation and the experiment do not match. Therefore, we create a modified method to improve the basic kinetic analysis. We can obtain the calibrated and in terms of this modified method. Using the calibrated and to simulate interaction curves, we can obtain more consistent simulation results with experiments.