Automation and Control of Micro-scale Biological Tasks: Single Cell Surgery

在生物科学中，过去十年见证了对单细胞生物学的兴趣日益增长，同时使这项工作成为可能，增加了工程研究中开发单细胞手术工具的兴趣。这些单细胞手术过程包括：（i）去除（活组织检查）细胞器，例如细胞核、线粒体，用于细胞器性质表征，（ii）注射化学标记物用于细胞内传感，（iii）将RNA、DNA转移到细胞中以产生转基因生物体，（iv）人类干细胞治疗中干细胞系的细胞组分活组织检查和（v）卵裂球或滋养外胚层活组织检查（人类胚胎细胞）植入前遗传学诊断（PGD），在儿童发育之前检测遗传疾病。在加拿大和世界各地的研究实验室和体外受精（IVF）诊所中，对这种过程的需求日益增加。目前，这些任务由高度熟练的技术人员使用显微镜和微量移液器进行。使用明视野显微镜，在其下细胞呈现透明。由于景深非常窄（小于几微米），只有细胞的一个非常薄的切片在焦点上。因此，在图像平面上方或下方的感兴趣的对象是极度失焦的。这种有限的焦点深度只有几微米。再加上操作者疲劳和潜在的人体污染，对细胞手术的生产量和成功率造成了显著的负面影响。这些问题的解决方案是通过将自动化制造原理应用于细胞手术任务，再加上实时3D图像反馈来控制自动化过程。消除人类操作员将导致增加的吞吐量和成功率。 ** 为了克服细胞手术中2D图像反馈的局限性，我们建议研究和开发一种用于细胞手术的实时3D成像系统，该系统将连续提取细胞手术所必需的3D信息，例如细胞器质心位置，细胞膜位置和其他量。 ** 研究计划的目标 * 这项拟议的研究旨在利用自动化机器人生物操作系统研究单细胞手术的自动化。*1。为机器人驱动的微量移液器自动化细胞手术开发自动化策略。* 2.开发基于3D视觉的方法，以促进细胞的定位和后续处理。 *3。在处理前对细胞进行细胞器鉴定。该成像将识别感兴趣的细胞器及其位置。*4。开发使用具有视觉反馈的MEMS设备的电磁方法，以在两个方向旋转中旋转悬浮细胞，用于成像和随后的细胞手术。 *5。开发基于机器人的控制方法，用于使用3D图像信息的自动细胞器活检和小细胞的细胞注射。*****

Within the biological sciences, the last decade has witnessed growing interest in single cell biology, and simultaneously enabling this work, increasing interest within engineering research to develop tools to carry out single cell surgery. These single cell surgery processes include: (i) removal (biopsy) of cell organelles e.g. nucleus, mitochondria, for organelle property characterization, (ii) injection of chemical markers for intracellular sensing, (iii) transfer of RNA, DNA into the cell to create transgenic organisms, (iv) biopsy of cell components for stem cell lines in human stem cell therapy and (v) blastomere or trophectoderm biopsy (human embryonic cell) Preimplantation Genetic Diagnosis (PGD) to detect genetic disease prior to the development of a child. Increasing demand for such processes is seen in both research laboratories and in-vitro fertilization (IVF) clinics in Canada and worldwide.******Currently, these tasks are carried out by highly skilled technicians, using microscopes, with micro-pipettes. Bright field microscopes are used, under which cells appear transparent. Due to the very narrow depth of field (less than a few microns), only a very thin slice of the cell is in focus. Hence, objects of interest above or below the image plane, are extremely out of focus. This limited depth of focus of only a few µm. coupled with operator fatigue and potential human contamination, contributes to a significant negative impact on throughput and success rate of cell surgery. A solution to these issues is found through the application of automated manufacturing principles to cell surgery tasks, coupled with real-time 3D image feedback to control the automation processes. Elimination of human operators will result in increased throughput and success rates. ******To overcome the limitations of 2D image feedback in cell surgery, we propose to investigate and develop a real-time 3D imaging system for cell surgery which will continuously extract 3D information essential for cell surgery, e.g. organelle centroid location, cell membrane location, and other quantities. ******Objectives of the Research Program***This proposed research aims to investigate automation of single cell surgery with an automated robotic bio-manipulation system. ***1. Develop automation strategies for robotic actuated micro-pipette automated cell surgery.***2. Develop 3D vision based methodologies to facilitate the location and subsequent processing of cells. ***3. Cell organelle identification will be carried out on cells prior to processing. This imaging will identify the cell organelle of interest, and its location. ***4. Develop electromagnetic approaches using MEMS device with vision feedback, to rotate suspended cells in two direction rotations, for imaging and subsequent cell surgery. ***5. Develop robotic based control methods for automated cell organelle biopsy and cell injection for small cells using 3D image information. *****