Visually-guided protein crystal manipulation using micromachined silicon tools

We present a system for protein crystal micro-manipulation with focus on automated crystal mounting for the purposes of X-ray data collection. The system features a set of newly designed micropositioner end-effectors we call microshovels which address some limitations of the traditional cryogenic loops. We have used micro-electrical mechanical system (MEMS) techniques to design and manufacture various shapes and quantities of microshovels. Visual feedback from a camera mounted on the microscope is used to control the micropositioner as it lowers a microshovel into the liquid containing the crystals and approaches a selected crystal for pickup. We present experimental results that illustrate the applicability of our approach

Visually-guided protein crystal manipulation using micromachined silicon tools

We present a system for protein crystal micro-manipulation with focus on automated crystal mounting for the purposes of X-ray data collection. The system features a set of newly designed micropositioner end-effectors we call microshovels which address some limitations of the traditional cryogenic loops. We have used micro-electrical mechanical system (MEMS) techniques to design and manufacture various shapes and quantities of microshovels. Visual feedback from a camera mounted on the microscope is used to control the micropositioner as it lowers a microshovel into the liquid containing the crystals and approaches a selected crystal for pickup. We present experimental results that illustrate the applicability of our approach

Evaluating the role of force feedback for biomanipulation tasks

Paper presented at the IEEE Virtual Reality, Haptics Symposium and Symposium on 3D User Interface, Alexandria, VA.Conventional cell manipulation techniques do not have the ability to provide force feedback to an operator. Poor control of cell injection force is one of the primary reasons for low success rates in cell injection and transgenesis in particular. Therefore, there exists a need to incorporate force feedback into a cell injection system. We have developed an automated cell injection system, which has the capability of measuring forces in the range of μN. We tested our system with 40 human subjects to evaluate the role of force feedback in cell injection task. Our experimental results indicate that the subjects were able to feel the cell injection force and confirmed our research hypothesis that the use of combined vision and force feedback leads to higher success rate in cell injection task compared to using vision feedback alone

Protein crystal presentation for synchrotron methods:acoustic techniques

This thesis describes the design, development and testing of novel acoustic mounting methods for the diffraction of protein crystals for structural biology. Sample environment and presentation is a challenging field set within the larger subject of protein crystallography. The needs of researchers to achieve the highest resolution data collection from difficult to fabricate samples, sits alongside the need for complex experimental conditions, where temperature, hydration and chemistry must be altered and controlled. A movement within the structural biology field away from obtaining structures at cryogenic temperatures, towards high resolution structures at room temperature, where proteins may still function, has been the driver to search for novel solutions. To approach this ‘solution space’ the following work draws on acoustic manipulation techniques, looking for self-assembly and non-contact manipulation. Thus for the first time acoustic standing wave crystal trapping and also acoustically induced rotation have been shown in situ to be viable for use with protein crystallography, a fundamental proof paving the way for new time resolved and high throughput methods. The methods investigated included both surface acoustic and bulk acoustic waves, looking at self-assembly and flow induced rotation. Each demonstration addresses a fundamental need within the automation of room temperature crystallography, demonstrating both the technique and quantifying the diffraction resolution for the first time. Through the completion of these novel experiments acoustic sample presentation has been proven viable. The demonstrated method does not require crystals be removed from crystallisation fluid before mounting (using the acoustic trapping method), thus enabling the application of secondary fluids and paving the way for a fully continuous and microfluidic technology. Moreover acoustic goniometry lends itself to the automated mounting and collection of small batch crystal data by removing the need for delicate spine mounting. Both methods constitute a significant extension in the ability of researchers to utilise non-contact methods to control and interact with their proteins and crystals at room temperature

Mechanical Manipulation and Characterization of Biological Cells

Mechanical manipulation and characterization of an individual biological cell is currently one of the most exciting research areas in the field of medical robotics. Single cell manipulation is an important process in intracytoplasmic sperm injection (ICSI), pro-nuclei DNA injection, gene therapy, and other biomedical areas. However, conventional cell manipulation requires long training and the success rate depends on the experience of the operator. The goal of this research is to address the drawbacks of conventional cell manipulation by using force and vision feedback for cell manipulation tasks. We hypothesize that force feedback plays an important role in cell manipulation and possibly helps in cell characterization. This dissertation will summarize our research on: 1) the development of force and vision feedback interface for cell manipulation, 2) human subject studies to evaluate the addition of force feedback for cell injection tasks, 3) the development of haptics-enabled atomic force microscope system for cell indentation tasks, 4) appropriate analytical model for characterizing the mechanical property of mouse embryonic stem cells (mESC) and 5) several indentation studies on mESC to determine the mechanical property of undifferentiated and early differentiating (6 days under differentiation conditions) mESC. Our experimental results on zebrafish egg cells show that a system with force feedback capability when combined with vision feedback can lead to potentially higher success rates in cell injection tasks. Using this information, we performed experiments on mESC using the AFM to understand their characteristics in the undifferentiated pluripotent state as well as early differentiating state. These experiments were done on both live as well as fixed cells to understand the correlation between the two during cell indentation studies. Our results show that the mechanical property of undifferentiated mESC differs from early differentiating (6th day) mESC in both live and fixed cells. Thus, we hypothesize that mechanical characterization studies will potentially pave the way for developing a high throughput system with force feedback capability, to understand and predict the differentiation path a particular pluripotent cell will follow. This finding could also be used to develop improved methods of targeted cellular differentiation of stem cells for therapeutic and regenerative medicine