Wang Han, Kim Jeongyun, Jayaraman Arul, Han Arum
Department of Electrical and Computer Engineering, Texas A&M University, College Station, TX, 77843, USA.
Biomed Microdevices. 2014 Dec;16(6):887-96. doi: 10.1007/s10544-014-9893-x.
Microfluidic live cell arrays with integrated concentration gradient or mixture generators have been utilized in screening cellular responses to various biomolecular cues. Microfluidic network-based gradient generators that can create concentration gradients by repeatedly splitting and mixing different solutions using networks of serpentine channels are commonly used. However, in this method the generation of concentration gradients relies on the continuous flow of sample solutions at optimized flow rates, which poses challenges in maintaining the pressure and flow stability throughout the entire assay period. Here we present a microfluidic live cell screening array with an on-demand multi-reagent mixture generator where the mixing ratios, thus generated concentrations, are hard-wired into the chip itself through a geometric metering method. This platform showed significantly improved robustness and repeatability in generating concentration gradients of fluorescent dyes (average coefficient of variance C.V. = 9 %) compared to the conventional network-based gradient generators (average C.V. = 21 %). In studying the concentration dependent effects of the environmental toxicant 3-methylcholanthrene (3MC) on the activation of cytochrome P450 1A1 (Cyp 1A1) enzyme in H4IIE rat hepatoma cells, statistical variation of the Cyp 1A1 response was significantly lower (C.V. = 5 %) when using the developed mixture generator compared to that using the conventional gradient generator (C.V. = 12 %). Reduction in reagent consumption by 12-times was also achieved. This robust, accurate, and scalable multi-reagent mixture generator integrated with a cell culture array as a live cell assay platform can be readily implemented into various screening applications where repeatability, robustness, and low reagent consumptions over long periods of assay time are of importance.
具有集成浓度梯度或混合物发生器的微流控活细胞阵列已被用于筛选细胞对各种生物分子信号的反应。基于微流控网络的梯度发生器通常通过使用蛇形通道网络反复分离和混合不同溶液来创建浓度梯度。然而，在这种方法中，浓度梯度的产生依赖于样品溶液以优化流速的连续流动，这在整个检测期间维持压力和流量稳定性方面带来了挑战。在此，我们展示了一种具有按需多试剂混合物发生器的微流控活细胞筛选阵列，其中通过几何计量方法将混合比例以及由此产生的浓度硬连线到芯片本身。与传统的基于网络的梯度发生器（平均变异系数C.V. = 21%）相比，该平台在生成荧光染料浓度梯度方面显示出显著提高的稳健性和可重复性（平均变异系数C.V. = 9%）。在研究环境毒物3-甲基胆蒽（3MC）对H4IIE大鼠肝癌细胞中细胞色素P450 1A1（Cyp 1A1）酶激活的浓度依赖性影响时，与使用传统梯度发生器（C.V. = 12%）相比，使用开发的混合物发生器时Cyp 1A1反应的统计变异性显著更低（C.V. = 5%）。试剂消耗也减少了12倍。这种稳健、准确且可扩展的多试剂混合物发生器与细胞培养阵列集成作为活细胞检测平台，可以很容易地应用于各种筛选应用中，在这些应用中，长时间检测的重复性、稳健性和低试剂消耗至关重要。
