Robotic Micromanipulation and Mechanical Characterization of Intracellular Structures

在过去的二十年里，单细胞操作和测量取得了长足的进步。这项研究将超越最先进的水平，开发一种用于细胞内3D操作和机械测量的变革性平台技术。新的平台和技术将使细胞内亚微米磁珠(以及后来更复杂的磁性设备)能够进行机械测量的3D导航，这将开启细胞内物理测量和操作的令人兴奋的新纪元。 近期目标包括：目标1：建立用于3D细胞内导航和测量的新的机器人磁钳平台；目标2：开发新的磁力模型和力/位置控制方法，用于利用3D共聚焦显微镜图像反馈控制亚微米磁珠；目标3：在完整细胞的细胞核上进行不同位置的力学测量，以量化沿细胞核不同方向的硬度异质性。为了实现这些目标，我们将开发一种新的多极磁钳平台和技术来操纵亚微米磁珠，并解决低带宽高分辨率视觉反馈、亚微米位置控制和皮牛顿力控制等挑战。具体来说，我们将致力于三维磁钳平台的设计、建造、分析建模、数值模拟、系统测试/调试和优化。我们将建立非线性磁场模型，将驱动电流与磁场产生联系起来，并将力的应用量化。还将进行磁场模拟，以计算施加在亚微米磁珠上的磁力。将进行标定实验，并将在实验结果和工作空间中每个点的解析建模和有限元仿真结果中量化位置力误差。 基于微珠动力学的广义预测控制器将被开发来将亚微米微珠定位到完整细胞的细胞核上的不同位置，以不同的速度压入细胞核。实验数据将用力学模型进行分析，以生成癌细胞发育不同阶段细胞核上每个测量位置的硬度/弹性/粘度图。将测量沿细胞核长轴和短轴的机械性能的极性，力诱导的软轴僵硬，以及癌细胞迁移行为的改变。细胞核的机械极性对于癌细胞迁移的必要性将被明确地证明。这项研究计划将培训HQP在NSE从事研究职业，或满足加拿大仪器和生物技术行业的巨大需求。

The past two decades have seen strides in single-cell manipulation and measurement. This research will go beyond the state of the art to develop a transformative platform technology for 3D manipulation and mechanical measurement inside a cell. The new platform and techniques will enable 3D navigation of a sub-micron magnetic bead (and later more sophisticated magnetic devices) inside a cell to perform mechanical measurements, which will open an exciting new era of intracellular physical measurement and manipulation. The near-term objectives include: Aim 1: Establish a new robotic magnetic tweezers platform for 3D intracellular navigation and measurement; Aim 2: Develop new magnetic force models and force/position control methods for controlling a sub-micron magnetic bead with 3D confocal microscopy image feedback; Aim 3: Perform mechanical measurements at different locations on the cell nucleus in an intact cell to quantify stiffness heterogeneity along different directions of the cell nucleus. To achieve these goals, we will develop a new multi-pole magnetic tweezers platform and technologies to manipulate a sub-micron magnetic bead and tackle the challenges of low-bandwidth high-resolution visual feedback, sub-micron position control, and picoNewton force control. Specifically, we will pursue 3D magnetic tweezers platform design, construction, analytical modeling, numerical simulation, system testing/debugging, and optimization. We will establish nonlinear magnetic field models that relate driving currents to magnetic field generation and quantify force application. Magnetic field simulation will also be conducted to calculate magnetic forces exerted on the sub-micron magnetic bead. Calibration experiments will be performed and position-force errors will be quantified among experimental results and results from analytical modeling and finite element simulation for each point in the workspace. Generalized predictive controllers based on bead dynamics will be developed to position a sub-micron bead to different locations on the cell nucleus in an intact cell to indent the cell nucleus at varying speeds. Experimental data will be analyzed with mechanics models to generate a stiffness/modulus/viscosity map for each measured location on the cell nucleus at different stages of cancer cell development. Polarity of mechanical properties along the major and minor axes of the cell nucleus, force-induced stiffening of the softer axis, and alteration of the migratory behavior of cancer cells will be measured. The necessity of mechanical polarity of the cell nucleus for cancer cell migration will be unequivocally demonstrated. This research program will train HQP to pursue research careers in NSE or fulfill the great demand of Canadian instrumentation and biotech industries.