Department of Aerospace and Mechanical Engineering, University of Notre Dame, Notre Dame, Indiana 46556, USA.
Lab Chip. 2015 Apr 7;15(7):1656-66. doi: 10.1039/c5lc00036j. Epub 2015 Feb 18.
There has been increasing evidence that micro and messenger RNA derived from exosomes play important roles in pancreatic and other cancers. In this work, a microfluidics-based approach to the analysis of exosomal RNA is presented based on surface acoustic wave (SAW) exosome lysis and ion-exchange nanomembrane RNA sensing performed in conjunction on two separate chips. Using microRNA hsa-miR-550 as a model target and raw cell media from pancreatic cancer cell lines as a biological sample, SAW-based exosome lysis is shown to have a lysis rate of 38%, and an ion-exchange nanomembrane sensor is shown to have a limit of detection of 2 pM, with two decades of linear dynamic range. A universal calibration curve was derived for the membrane sensor and used to detect the target at a concentration of 13 pM in a SAW-lysed sample, which translates to 14 target miRNA per exosome from the raw cell media. At a total analysis time of 1.5 h, this approach is a significant improvement over existing methods that require two overnight steps and 13 h of processing time. The platform also requires much smaller sample volumes than existing technology (100 μL as opposed to ~mL) and operates with minimal sample loss, a distinct advantage for studies involving mouse models or other situations where the working fluid is scarce.
越来越多的证据表明，来源于外泌体的微小 RNA 和信使 RNA 在胰腺和其他癌症中发挥着重要作用。在这项工作中，提出了一种基于微流控的外泌体 RNA 分析方法，该方法基于表面声波 (SAW) 外泌体裂解和在两个单独的芯片上联合进行的离子交换纳米膜 RNA 传感。使用 microRNA hsa-miR-550 作为模型靶标，并用胰腺癌细胞系的原始细胞培养基作为生物样品，结果表明，基于 SAW 的外泌体裂解的裂解率为 38%，离子交换纳米膜传感器的检测限为 2 pM，线性动态范围为两个数量级。为膜传感器推导了通用校准曲线，并用于检测在 SAW 裂解样品中浓度为 13 pM 的靶标，这相当于原始细胞培养基中外泌体中 14 个靶标 miRNA。总分析时间约为 1.5 小时，与需要两个过夜步骤和 13 小时处理时间的现有方法相比，该方法有了显著的改进。该平台还需要比现有技术小得多的样品体积（100 μL 而不是mL），并且几乎没有样品损失，这对于涉及小鼠模型或其他工作流体稀缺的情况的研究是一个明显的优势。
