Park Juhwan, Roh Hyewon, Park Je-Kyun
Department of Bio and Brain Engineering, Korea Advanced Institute of Science and Technology (KAIST), 291 Daehak-ro, Yuseong-gu, Daejeon 34141, Korea.
Micromachines (Basel). 2019 Mar 2;10(3):174. doi: 10.3390/mi10030174.
The generation of concentration gradients is an essential part of a wide range of laboratory settings. However, the task usually requires tedious and repetitive steps and it is difficult to generate concentration gradients at once. Here, we present a microfluidic device that easily generates a concentration gradient by means of push-button actuated pumping units. The device is designed to generate six concentrations with a linear gradient between two different sample solutions. The microfluidic concentration gradient generator we report here does not require external pumps because changes in the pressure of the fluidic channel induced by finger actuation generate a constant volume of fluid, and the design of the generator is compatible with the commonly used 96-well microplate. Generation of a concentration gradient by the finger-actuated microfluidic device was consistent with that of the manual pipetting method. In addition, the amount of fluid dispensed from each outlet was constant when the button was pressed, and the volume of fluid increased linearly with respect to the number of pushing times. Coefficient of variation (CV) was between 0.796% and 13.539%, and the error was between 0.111% and 19.147%. The design of the microfluidic network, as well as the amount of fluid dispensed from each outlet at a single finger actuation, can be adjusted to the user's demand. To prove the applicability of the concentration gradient generator, an enzyme assay was performed using alkaline phosphatase (ALP) and -nitrophenyl phosphate (NPP). We generated a linear concentration gradient of the NPP substrate, and the enzyme kinetics of ALP was studied by examining the initial reaction rate between ALP and NPP. Then, a Hanes⁻Woolf plot of the various concentration of ALP was drawn and the and value were calculated.
浓度梯度的产生是众多实验室操作的重要组成部分。然而，这项任务通常需要繁琐且重复的步骤，并且很难一次性产生浓度梯度。在此，我们展示一种微流控装置，它通过按钮驱动的泵送单元能够轻松产生浓度梯度。该装置旨在在两种不同的样品溶液之间产生具有线性梯度的六种浓度。我们在此报告的微流控浓度梯度发生器不需要外部泵，因为手指驱动引起的流体通道压力变化会产生恒定体积的流体，并且该发生器的设计与常用的96孔微孔板兼容。通过手指驱动的微流控装置产生的浓度梯度与手动移液法一致。此外，按下按钮时，每个出口分配的流体量是恒定的，并且流体体积相对于按压次数呈线性增加。变异系数（CV）在0.796%至13.539%之间，误差在0.111%至19.147%之间。微流控网络的设计以及单次手指驱动时每个出口分配的流体量可以根据用户需求进行调整。为了证明浓度梯度发生器的适用性，使用碱性磷酸酶（ALP）和对硝基苯磷酸酯（NPP）进行了酶测定。我们生成了NPP底物的线性浓度梯度，并通过检测ALP与NPP之间的初始反应速率来研究ALP的酶动力学。然后，绘制了不同浓度ALP的Hanes-Woolf图，并计算了米氏常数和最大反应速度。
