Three-dimensional microscope tracking system using the astigmatic lens method and a profile sensor

We developed a three-dimensional microscope tracking system using the astigmatic lens method and a profile sensor, which provides three-dimensional position detection over a wide range at the rate of 3.2 kHz. First, we confirmed the range of target detection of the developed system, where the range of target detection was shown to be +/- 90 mu m in the horizontal plane and +/- 9 mu m in the vertical plane for a 10x objective lens. Next, we attempted to track a motion-controlled target. The developed system kept the target at the center of the field of view and in focus up to a target speed of 50 mu m/s for a 20x objective lens. Finally, we tracked a freely moving target. We successfully demonstrated the tracking of a 10-mu m-diameter polystyrene bead suspended in water for 40 min. The target was kept in the range of approximately 4.9 mu m around the center of the field of view. In addition, the vertical direction was maintained in the range of +/- 0.84 mu m, which was sufficiently within the depth of focus.