Faculty of Biomedical Engineering , Technion-Israel Institute of Technology , Haifa , 3200003 Israel.
Department of Physiology, Biophysics and Systems Biology, Faculty of Medicine , Technion , Haifa , 3109601 Israel.
Anal Chem. 2018 Jun 19;90(12):7480-7488. doi: 10.1021/acs.analchem.8b01017. Epub 2018 May 30.
A multitude of cell screening assays for diagnostic and research applications rely on quantitative measurements of a sample in the presence of different reagent concentrations. Standard methods rely on microtiter plates of varying well density, which provide simple and standardized sample addressability. However, testing hundreds of chemical dilutions requires complex automation, and typical well volumes of microtiter plates are incompatible with the analysis of a small number of cells. Here, we present a microfluidic device for creating a high-resolution chemical gradient spanning 200 nanoliter wells. Using air-based shearing, we show that the individual wells can be compartmentalized without altering the concentration gradient, resulting in a large set of isolated nanoliter cell culture wells. We provide an analytical and numerical model for predicting the concentration within each culture chamber and validate it against experimental results. We apply our system for the investigation of yeast cell metabolic gene regulation in the presence of different ratios of galactose/glucose concentrations and successfully resolve the nutrient threshold at which the cells activate the galactose pathway.
许多用于诊断和研究应用的细胞筛选检测依赖于在不同试剂浓度存在下对样品进行定量测量。标准方法依赖于具有不同孔密度的微量滴定板，它提供了简单且标准化的样品可寻址性。然而，测试数百种化学稀释液需要复杂的自动化，而典型的微量滴定板孔体积与少量细胞的分析不兼容。在这里，我们提出了一种用于创建跨越 200 纳升孔的高分辨率化学梯度的微流控装置。使用基于空气的剪切，我们证明可以在不改变浓度梯度的情况下将各个孔分隔开，从而产生了大量的隔离的纳升细胞培养孔。我们提供了一个用于预测每个培养腔内浓度的分析和数值模型，并通过实验结果对其进行了验证。我们将我们的系统应用于研究酵母细胞在不同半乳糖/葡萄糖浓度比存在下的代谢基因调控，并成功确定了细胞激活半乳糖途径的营养阈值。
