Department of Agricultural and Biosystems Engineering, The University of Arizona, Tucson, AZ 85721-0038, USA.
J Biol Eng. 2012 Sep 4;6(1):15. doi: 10.1186/1754-1611-6-15.
A computer numerical control (CNC) apparatus was used to perform droplet centrifugation, droplet DNA extraction, and rapid droplet thermocycling on a single superhydrophobic surface and a multi-chambered PCB heater. Droplets were manipulated using "wire-guided" method (a pipette tip was used in this study). This methodology can be easily adapted to existing commercial robotic pipetting system, while demonstrated added capabilities such as vibrational mixing, high-speed centrifuging of droplets, simple DNA extraction utilizing the hydrophobicity difference between the tip and the superhydrophobic surface, and rapid thermocycling with a moving droplet, all with wire-guided droplet manipulations on a superhydrophobic surface and a multi-chambered PCB heater (i.e., not on a 96-well plate). Serial dilutions were demonstrated for diluting sample matrix. Centrifuging was demonstrated by rotating a 10 μL droplet at 2300 round per minute, concentrating E. coli by more than 3-fold within 3 min. DNA extraction was demonstrated from E. coli sample utilizing the disposable pipette tip to cleverly attract the extracted DNA from the droplet residing on a superhydrophobic surface, which took less than 10 min. Following extraction, the 1500 bp sequence of Peptidase D from E. coli was amplified using rapid droplet thermocycling, which took 10 min for 30 cycles. The total assay time was 23 min, including droplet centrifugation, droplet DNA extraction and rapid droplet thermocycling. Evaporation from of 10 μL droplets was not significant during these procedures, since the longest time exposure to air and the vibrations was less than 5 min (during DNA extraction). The results of these sequentially executed processes were analyzed using gel electrophoresis. Thus, this work demonstrates the adaptability of the system to replace many common laboratory tasks on a single platform (through re-programmability), in rapid succession (using droplets), and with a high level of accuracy and automation.
采用计算机数控（CNC）设备在单个超疏水表面和多腔印刷电路板（PCB）加热器上进行液滴离心、液滴 DNA 提取和快速液滴热循环。使用“线引导”方法（在本研究中使用移液器吸头）操纵液滴。该方法可以轻松适应现有的商业机器人移液系统，同时还展示了附加功能，例如振动混合、液滴高速离心、利用吸头和超疏水表面之间的疏水性差异进行简单的 DNA 提取，以及使用移动液滴进行快速热循环，所有这些都可以在超疏水表面和多腔 PCB 加热器上进行（即不在 96 孔板上）进行线引导的液滴操纵。还演示了用于稀释样品基质的系列稀释。通过以 2300 转/分钟的速度旋转 10μL 液滴来实现离心，在 3 分钟内将大肠杆菌浓缩超过 3 倍。从大肠杆菌样品中提取 DNA 时，利用一次性移液器吸头巧妙地将提取的 DNA 从位于超疏水表面的液滴中吸引出来，整个过程耗时不到 10 分钟。提取后，使用快速液滴热循环扩增来自大肠杆菌的 Peptidase D 的 1500bp 序列，30 个循环耗时 10 分钟。总测定时间为 23 分钟，包括液滴离心、液滴 DNA 提取和快速液滴热循环。在这些过程中，10μL 液滴的蒸发并不明显，因为最长的空气暴露时间和振动时间都不到 5 分钟（在 DNA 提取过程中）。使用凝胶电泳分析这些顺序执行过程的结果。因此，该工作证明了该系统能够在单个平台上（通过可重新编程性）快速连续（使用液滴）执行许多常见的实验室任务，并且具有高度的准确性和自动化程度。
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