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RoboLase : A robotic laser scissors and laser tweezers microscope Linda Z. Shi*, Jaclyn M. Nascimento, Nicole Wakida, Alexander Dvornikov, Norman Baker, Elliot L. Botvinick and Michael W. Berns Abstract-We have built a robotic laser scissors and laser tweezers microscope ( RoboLase ) that can be operated via the internet. The system can be used to image, ablate, and/or trap cells and their organelles by remote-control. In the ablation mode, RoboLase is being used to perform delicate microsurgery on mitotic organelles (individual microtubules, spindle fibers, and centrioles). In the trapping mode, the system is being used as a real-time automatic tracking and trapping system (RATTS) of fast moving cells. RATTS performs all tracking and trapping functions without human intervention and has been used remotely between Australia and the US. Index Terms-laser microscope scissors, laser tweezers, robotic I. INTRODUCTION Technology is revolutionizing the biomedical field with the latest development of automatic image processing algorithms and real-time robotic devices. Automated image processing algorithms have been successfully applied to tracking neurons (1-2), Caenorhabditis elegans (3), and sperm cells (4-5), identifying Sphacelaria algae (6) and soil bacteria (7), and reconstructing live embryos (8). Robotic telemicroscopy has been developed for general applications in (9-10) and is currently being applied to the pathology (11- 13) and microsurgery (14-15). Lasers are useful tools for micromanipulation of biological specimens (16). With the addition of tightly focused laser beams, microscopes have been turned into elaborate preparative tools that not only allow detailed observation of a specimen but also the capture, displacement, and microdissection of biological samples in vitro with astonishing ease and accuracy (17). Some commercial optical trapping and scissors systems are available, such as The LaserTweezers R Workstation and The LaserScissors*R Workstation from Cell Robotics Inc, and PALM MicroLaser Systems from P.A.L.M. Manuscript received October 29, 2006. This work was supported in part by the Air force Office of Scientific Research (AFOSR # F9620-00-1-0371) to MWB, and the Beckman Laser Institute Foundation. M. W. Berns is with Beckman Laser Institute and Department of Biomedical Engineering, University of California, Irvine, (phone: 949-824- 7565; fax: (714) 824-8413; e-mail: mwbernsguci.edu). L. Z. Shi, J. M. Nasc mento, B. a ker, are wltl University of Califoria, San Diego. Nico e Wakida, Alexander Dvorn kov, Elliot Botvinick are with the Beckman Laser Institute University of California, Irvine. Microlaser Technologies. Consistent with the need to develop sophisticated new nanosystem technologies, we have built a robotic laser scissors and laser tweezers microscope ( RoboLase ) that can be remotely operated using either internet or dedicated line technology. The system can be used to image, ablate, and/or trap moving cells and their organelles. For the laser ablation application (Figure 1), the microscope stage, microscope parameters (reflector turret, light path, light intensity, objective selection and focus control), laser and arc lamp shutters. laser power, laser fire position, camera parameters and camera control are all controlled by the software. The user can draw multiple different shapes (dot, line, rectangle, circle. irregular shapes) on the captured cell image using the computer mouse. A click of a button on the computer screen instructs the laser to outline or cut out the shapes at the pre-set laser power. For the sequence acquiring image mode, the stage can be moved either by a joystick or by the computer mouse, and the objective focus can be adjusted by a mouse-click. For the time series mode, the phase or fluorescence images can be captured in a fixed-time interval and saved to the hard disk. The time series can also be paused if the interested cell is swimming out of focus. The histogram of the image is displayed on the computer screen to aid the user in adjusting the image display intensity by varying the maximum and minimum intensity values. For the laser trapping application, phase contrast images of swimming cells are digitized to the computer at a video rate (30 fps). The custom algorithm creates a region of interest (ROI) centered about a cell in response to a mouse click and performs all subsequent tasks automatically. Microscope stage movement responds to feedback from video analysis of the swimming cell to center the cell with respect to the field of view. The cell is automatically relocated to the laser trap focus where it is trapped either at a fixed power for a user-defined duration, or at a gradually decreasing laser power until the cell escapes. The cell's position is automatically monitored in order to measure the laser power at which the cell escapes the trap. II. ROBOTIC LASER ABLATION MICROSCOPE SYSTEM A. System setup The detailed hardware setup of the robotic laser ablation microscope was described in (10), (18) and (19). Nearly

"RoboLase": Internet-Accessible Robotic Laser Scissors and Laser Tweezers Microscope Systems