Son Kyung Jin, Rahimian Ali, Shin Dong-Sik, Siltanen Christian, Patel Tushar, Revzin Alexander
Department of Biomedical Engineering, University of California, Davis, California 95616, USA.
Department of Cancer Biology, Mayo Clinic, Jacksonville, Florida 32224, USA.
Analyst. 2016 Jan 21;141(2):679-88. doi: 10.1039/c5an01648g.
Monitoring activity of single cells has high significance for basic science and diagnostic applications. Here we describe a reconfigurable microfluidic device for confining single cells along with antibody-modified sensing beads inside 20 picoliter (pL) microcompartments for monitoring cellular secretory activity. An array of ∼7000 microchambers fabricated in the roof of the reconfigurable microfluidic device could be raised or lowered by applying negative pressure. The floor of the device was micropatterned to contain cell attachment sites in registration with the microcompartments. Using this set-up, we demonstrated the detection of inflammatory cytokine IFN-γ and exosomes from single immune cells and cancer cells respectively. The detection scheme was similar in both cases: cells were first captured on the surface inside the microfluidic device, then sensing microbeads were introduced into the device so that, once the microcompartments were lowered, single cells and microbeads became confined together. The liquid bathing the beads and the cells inside the compartments also contained fluorescently-labeled secondary antibodies (Abs). The capture of cell-secreted molecules onto microbeads was followed by binding of secondary antibodies - this caused microbeads to become fluorescent. The fluorescence intensity of the microbeads changed over time, providing dynamics of single cell secretory activity. The microdevice described here may be particularly useful in the cases where panning upstream of sensing is required or to analyze secretory activity of anchorage-dependent cells.
监测单细胞的活性对于基础科学和诊断应用具有重要意义。在此，我们描述了一种可重构微流控装置，用于将单细胞与抗体修饰的传感微珠限制在20皮升（pL）的微隔室内，以监测细胞的分泌活性。通过施加负压，可重构微流控装置顶部制造的约7000个微腔阵列可以升高或降低。该装置的底部进行了微图案化处理，以包含与微隔室对齐的细胞附着位点。利用这种设置，我们分别展示了对来自单个免疫细胞和癌细胞的炎性细胞因子IFN-γ和外泌体的检测。两种情况下的检测方案相似：首先将细胞捕获在微流控装置内部的表面上，然后将传感微珠引入装置中，这样，一旦微隔室下降，单细胞和微珠就会被限制在一起。隔室内浸泡微珠和细胞的液体中还含有荧光标记的二抗（Abs）。细胞分泌的分子被捕获到微珠上后，二抗会与之结合，这会使微珠发出荧光。微珠的荧光强度随时间变化，从而提供单细胞分泌活性的动态信息。本文所述的微装置在需要在传感上游进行淘选或分析贴壁依赖性细胞的分泌活性的情况下可能特别有用。
