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

在生物科学领域，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，对开发进行单细胞手术的工具的工程研究的兴趣日益增加。这些单细胞手术过程包括：(i)移除（活检）细胞器，如细胞核、线粒体，以表征细胞器特性；（ii）注射用于细胞内传感的化学标记物；（iii）将RNA、DNA转移到细胞中以产生转基因生物；（iv）在人类干细胞治疗中对干细胞系进行细胞成分活检；(v)卵裂球或滋养外胚层活检（人类胚胎细胞）植入前遗传学诊断（PGD），以在儿童发育之前检测遗传病。在加拿大和世界各地的研究实验室和体外受精（IVF）诊所，对这种过程的需求不断增加。******目前，这些任务是由高技能的技术人员进行的，使用显微镜和微移液器。使用亮场显微镜，在这种显微镜下细胞看起来是透明的。由于非常窄的景深（小于几微米），只有一个非常薄的细胞片是在焦点上。因此，在图像平面以上或以下的感兴趣的物体都是极度失焦的。这种有限的聚焦深度只有几微米，再加上操作人员的疲劳和潜在的人为污染，对细胞手术的吞吐量和成功率产生了重大的负面影响。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，找到了解决这些问题的方法。消除人工操作将提高吞吐量和成功率。******为了克服二维图像反馈在细胞手术中的局限性，我们建议研究和开发一种用于细胞手术的实时三维成像系统，该系统可以连续提取细胞手术所需的三维信息，如细胞器质心位置、细胞膜位置等。******研究计划目标***本研究计划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。机器人驱动微移液自动细胞手术的自动化策略研究。***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。细胞器鉴定将在处理前对细胞进行。这种成像将确定感兴趣的细胞器及其位置。* * * 4。使用带有视觉反馈的MEMS设备开发电磁方法，以两个方向旋转悬浮细胞，用于成像和随后的细胞手术。* * * 5。利用3D图像信息开发基于机器人的自动化细胞器活检和小细胞注射的控制方法。＊＊＊＊＊

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. *****