The MOE Key Laboratory of Spectrochemical Analysis and Instrumentation, the Key Laboratory of Chemical Biology of Fujian Province, State Key Laboratory of Physical Chemistry of Solid Surfaces, Collaborative Innovation Center of Chemistry for Energy Materials, Department of Chemical Biology, College of Chemistry and Chemical Engineering, Pen-Tung Sah Institute of Micro-Nano Science and Technology, Xiamen University , Xiamen 361005, People's Republic of China.
The MOE Key Laboratory of Analysis and Detection Technology for Food Safety, State Key Laboratory of Photocatalysis on Energy and Environment, College of Biological Science and Engineering, Fuzhou University , Fuzhou 350116, People's Republic of China.
Anal Chem. 2018 Feb 20;90(4):2570-2577. doi: 10.1021/acs.analchem.7b04040. Epub 2018 Feb 6.
Compartmentalization of aqueous samples in uniform emulsion droplets has proven to be a useful tool for many chemical, biological, and biomedical applications. Herein, we introduce an array-based emulsification method for rapid and easy generation of monodisperse agarose-in-oil droplets in a PDMS microwell array. The microwells are filled with agarose solution, and subsequent addition of hot oil results in immediate formation of agarose droplets due to the surface-tension of the liquid solution. Because droplet size is determined solely by the array unit dimensions, uniform droplets with preselectable diameters ranging from 20 to 100 μm can be produced with relative standard deviations less than 3.5%. The array-based droplet generation method was used to perform digital PCR for absolute DNA quantitation. The array-based droplet isolation and sol-gel switching property of agarose enable formation of stable beads by chilling the droplet array at -20 °C, thus, maintaining the monoclonality of each droplet and facilitating the selective retrieval of desired droplets. The monoclonality of droplets was demonstrated by DNA sequencing and FACS analysis, suggesting the robustness and flexibility of the approach for single molecule amplification and analysis. We believe our approach will lead to new possibilities for a great variety of applications, such as single-cell gene expression studies, aptamer selection, and oligonucleotide analysis.
将水性样品分隔在均匀的乳液液滴中已被证明是许多化学、生物和生物医学应用的有用工具。在此，我们介绍了一种基于阵列的乳化方法，用于在 PDMS 微井阵列中快速轻松地生成单分散琼脂糖油滴。微井充满了琼脂糖溶液，随后加入热油会由于液体溶液的表面张力而立即形成琼脂糖液滴。由于液滴尺寸仅由阵列单元尺寸决定，因此可以使用相对标准偏差小于 3.5%的预选择直径为 20 至 100 μm 的均匀液滴。基于阵列的液滴生成方法用于进行绝对 DNA 定量的数字 PCR。琼脂糖的基于阵列的液滴隔离和溶胶-凝胶转换特性可通过将液滴阵列冷却至-20°C 来形成稳定的珠粒，从而保持每个液滴的单克隆性，并便于选择性回收所需的液滴。通过 DNA 测序和 FACS 分析证明了液滴的单克隆性，表明该方法用于单分子扩增和分析的稳健性和灵活性。我们相信，我们的方法将为各种应用带来新的可能性，例如单细胞基因表达研究、适体选择和寡核苷酸分析。
