Microfluidic Monitoring of Single Cell Elasticity, Viscoelasticity, and Plasticity

 描述（由申请人提供）：单个人细胞的机械刚度可能是揭示细胞功能障碍的关键参数。例如，已知疟疾感染的红细胞比未感染的细胞更硬，并且侵袭性癌细胞的变形性比健康表型高出数倍。然而，对于生物物理特性在生物医学和诊断环境中更有用，我们将需要高通量连续生物力学分级的方法，类似于尺寸排阻色谱法。虽然通常采用通过尺寸和密度等参数进行分离，但很少有方法可用于通过刚度进行高通量分离，也没有方法用于通过粘弹性进行分选。 此外，由于不同细胞类型的生物物理特征的潜在重叠，为了在分离中实现纯度，可能需要破碎细胞的能力，使得可以收集生物物理值不重叠的亚群。为此，我们创造了一种微流体分选技术，该技术利用流体动力学和压缩力的组合来通过生物物理特性分选单个细胞。本研究提案的目的是创建一种基于微流体技术的高通量细胞分级方法，该方法对细胞的两种正交生物物理表型-刚度和粘度敏感。该技术包括一个具有周期性对角收缩的微通道，当细胞流动时，这些收缩使细胞变形，从而按比例改变其轨迹， 细胞硬度和粘度。例如，较硬的细胞朝向通道的上部平移，较软的细胞朝向通道的底部迁移，使得出口可以连续地收集分选的细胞。通过设计通道几何形状，例如脊间间距，可以强调细胞的粘弹性松弛，构成了以前未使用的全新分选机制。通过对通道流体动力学和细胞的刚度依赖性轨迹的计算理解，可以设计出口以专门收集分选的细胞，从而通过刚度使细胞破碎。此外，通过设计具有不同脊间间距的通道，可以利用细胞弛豫速率的差异。 在初步的数据中，我们显示了超过45倍的细胞类型的富集是可能的，在一个无标记的方式。

 DESCRIPTION (provided by applicant): The mechanical stiffness of individual human cells can be a key parameter that reveals dysfunction of the cell. For example, malaria-infected red blood cells are known to be stiffer than uninfected cells and invasive cancer cells can be several times more deformable than healthy phenotypes. However, for biophysical properties to be more useful in biomedical and diagnostic settings, we will require methods for continuous biomechanical fractionation in high throughput, akin to size exclusion chromatography. Although separation by such parameters as size and density are commonly employed, few methods are available for high throughput separation by stiffness and no method for sorting by viscoelasticity. Moreover, because of the potential overlap of biophysical signatures of different cell types, to achieve purity in the separation may require the ability to fractionate cells such that subpopulations can be collected for which the biophysical values do not overlap. Towards these ends, we have created a microfluidic sorting technology that utilizes a combination of hydrodynamic and compressive forces to sort individual cells by biophysical properties. The objective of this research proposal is to create a high-throughput cell fractionation method based on the microfluidic technology that is sensitive to stiffness and viscosity, two orthogonal biophysical phenotypes of cells. The technology consists of a microchannel with periodical, diagonal constrictions that deform cells as they flow to modify their trajectory in a proportion to cell stiffness and viscosity. For example, cells that are stiffer are translated towards the upper part of the channel and cells that are softer migrate towards the bottom part of the channel such that outlets can continuously collect the sorted cells. By engineering channel geometry such as inter-ridge spacing, viscoelastic relaxation of cells can be emphasized, constituting a completely new sorting mechanism not previously utilized. Through computational understanding of channel hydrodynamics and stiffness-dependent trajectories of cells, outlets can be designed to specifically collect the sorted cells and thereby fractionate cells by stiffness. Also, by designin channels with different inter-ridge spacing, differences in cell relaxation rates can be exploited. In preliminary data, we show over 45-fold enrichment of cell types is possible in a label-free manner.