結腸腫瘤藥物高通量篩選之微流體裝置研發

Abstract

癌症藥物篩選中的藥物稀釋過程往往複雜且耗時，傳統的人工稀釋方法不僅勞動密集且易受稀釋誤差累積的影響。本研究開發了一個藥物高通量篩選之微流體裝置，應用於結腸腫瘤藥物篩選，可以簡易、快速並精準生成特定之藥物濃度梯度。微流體裝置以流體在低雷諾數條件下呈現的層流特性，利用 Hagen-Poiseuille 方程式進行設計，透過改變梯度流道中混合點兩側的流道長度比例，調配兩流體混合的體積比，實現了對濃度梯度生成的精確控制且克服流速變化對擴散混合的限制，加速藥物篩選流程。實驗中，我們使用牛血清白蛋白溶液 (Bovine Serum Albumin，BSA) 作為模擬實際藥品稀釋的樣品，透過布拉德福蛋白質定量法 (Bradford protein assay) 分析微流體裝置產生之稀釋濃度值，以此來驗證微流體裝置的功能性。最終，利用微流體裝置稀釋化療藥物 (Oxaliplatin) 產生多個藥物濃度，進行人類結腸腫瘤細胞 (HCT-116) 的藥物毒性測試。結果顯示，微流體裝置可以在高入口注入流率條件下，於30秒內快速達到穩定狀態，並精確生成濃度梯度及控制輸出至96孔盤之體積量。微流體裝置可精準生成 1/3×～1/300× 之稀釋倍率，其生成之 BSA 濃度梯度與目標之濃度最大誤差小於 6%，輸出至各個孔洞間的液體體積誤差於 5% 以內。在結腸腫瘤藥物毒性測試中，透過微流體裝置稀釋方法與受過專業訓練的研究人員進行人工稀釋方法所得到的 IC50 誤差值僅為 2.45%。本研究開發的藥物高通量篩選微流體裝置可以在短時間內精準地生成目標之濃度，顯著提升了藥物篩選的效率和操作的便利性，在未來可結合細胞晶片技術，進一步研究藥物對特定病理狀態細胞的影響，推動個體化醫療和精準治療的發展。

In cancer drug screening, the dilution process of drugs is often complex and time-consuming. Traditional manual dilution methods are labor-intensive and susceptible to cumulative dilution errors. This study develops a microfluidic device for colorectal cancer drug high-throughput screening. The device simplifies, accelerates, and accurately generates specific drug concentration gradients. The microfluidic device leverages the laminar flow characteristics of fluids under low Reynolds number conditions and is designed using the Hagen-Poiseuille equation. The volumetric ratio of the two mixing fluids can be adjusted by varying the channel length ratio on both sides of the merging points in the gradient channel, achieving precise control over the concentration gradient generation. This design overcomes the limitations of diffusion mixing due to flow rate variations, thus expediting the drug screening process. The experiments used bovine serum albumin (BSA) solution as a sample to simulate actual drug dilution. The dilution concentrations produced by the microfluidic device were analyzed using the Bradford protein assay to validate its functionality. Ultimately, the microfluidic device was used to dilute the chemotherapy drug Oxaliplatin to produce multiple drug concentrations for cytotoxicity testing on human colon tumor cells (HCT-116). The results showed that the microfluidic device could rapidly reach a stable state within 30 seconds under high inlet flow rates. The device could accurately generate dilution ratios ranging from 1/3× to 1/300×, with the generated BSA concentration gradient having a maximum deviation of less than 6% from the target concentration and the volume deviation of the liquid output to each well within 5%. In the colorectal cancer drug cytotoxicity test, the deviation of IC50 between the microfluidic device dilution method and the manual dilution method performed by professionally trained researchers was only 2.45%. The microfluidic device developed in this study can accurately generate the target concentrations quickly, significantly enhancing the efficiency and convenience of drug screening. In the future, this device can be integrated with cell chip technology to further study drugs' effects on cells in specific pathological states, promoting the development of personalized medicine and precision therapy.