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 µm in the horizontal plane and ±?9 µm in the vertical plane for a 10× 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 µm/s for a 20× objective lens. Finally, we tracked a freely moving target. We successfully demonstrated the tracking of a 10-µm-diameter polystyrene bead suspended in water for 40 min. The target was kept in the range of approximately 4.9 µm around the center of the field of view. In addition, the vertical direction was maintained in the range of ±?0.84 µm, which was sufficiently within the depth of focus.