Low-stress Microfluidic Density-gradient Centrifugation for Blood Cell Sorting
Low-stress Microfluidic Density-gradient Centrifugation for Blood Cell Sorting
复制标题
DOI：
10.1007/s10544-018-0323-3
复制
发表时间：
2018-08-28
影响因子：
2.8
通讯作者：
Sethu, Palaniappan
中科院分区：
文献类型：
--
作者：
Sun, Yuxi;Sethu, Palaniappan
密度梯度离心利用不同血细胞之间的密度差异来实现外周血单核细胞（PBMC）与多形核（PNM）细胞和红细胞或红细胞（RBC）的分离。虽然密度梯度离心提供了一种无标记的替代方法，避免了使用苛刻的裂解缓冲液进行血细胞分离，但这是一种耗时且劳动密集型的过程，在此过程中，血细胞受到高水平的离心力，这可能会人为地激活细胞。为了提供这种优雅方法的低应力替代方案，我们使用微流体技术使该过程小型化和自动化，以确保从全血中连续分离PBMC，同时避免以简单的流通形式暴露于高水平的离心应力。在该装置内，通过利用微流体通道内的层流来建立密度梯度，以在较大的Ficoll流上分层薄的血流。使用这种方法，我们证明了成功的分离PBMC从全血中保存的单核细胞和不同的淋巴细胞亚群类似于传统的密度梯度离心。使用该技术分离的PBMC的活化状态的评价表明，与常规裂解或密度梯度离心相比，我们的方法实现了最小的分离过程诱导的细胞活化。这种简单、自动化的微流体密度梯度离心技术可以潜在地用作快速和无活化技术的工具，用于从全血中分离PBMC以用于护理点应用。
Density gradient centrifugation exploits density differences between different blood cells to accomplish separation of peripheral blood mononuclear cells (PBMCs) from polymorphonuclear (PNM) cells, and erythrocytes or red blood cells (RBCs). While density gradient centrifugation offers a label-free alternative avoiding the use of harsh lysis buffers for blood cell isolation, it is a time-consuming and labor-intensive process during which blood cells are subject to high-levels of centrifugal force that can artifactually activate cells. To provide a low-stress alternative to this elegant method, we miniaturized and automated this process using microfluidics to ensure continuous PBMCs isolation from whole blood while avoiding the exposure to high-levels of centrifugal stress in a simple flow-through format. Within this device, a density gradient is established by exploiting laminar flow within microfluidic channels to layer a thin stream of blood over a larger stream of Ficoll. Using this approach we demonstrate successful isolation of PBMCs from whole blood with preservation of monocytes and different lymphocyte subpopulations similar to that seen with conventional density gradient centrifugation. Evaluation of activation status of PBMCs isolated using this technique shows that our approach achieves minimal isolation process induced activation of cells in comparison to conventional lysis or density gradient centrifugation. This simple, automated microfluidic density gradient centrifugation technique can potentially serve as tool for rapid and activation-free technique for isolation of PBMCs from whole blood for point-of-care applications.