A design of a microscopy system tailored for optical tweezers with a capability of an automatic focusing is presented. In this design, we utilize lenses, motorized mechanical stage, lamp and a digital camera to magnify and see a micrometer sized spheres floating in a thin film of water. The system can automatically translate the stage that holds the specimen to obtain the best focused image. The best focused image is "sharp." Mathematically, the best focused image shows the maximum amount of high frequency terms from the images obtained by translating the stage. The metric that calculates how one image is focused is called the Focus Measure (FM). Unfortunately, low frequency components also increase this FM. And an optical imaging system is a low pass filtering system. Thus the primary concern is to lower the low frequency components in an image. The electric signals from each pixel of a CCD include noises that are inherent in each pixel. The result of this is an FM profile that has multiple local maxima. This is the most critical reason why an Automatic Focusing System (AFS) yields incorrect focusing results. Available techniques have been tested and from this experience, the most appropriate Focus Measure Filter (FMF) that has the sharpest FM despite the low frequency terms and noises has been selected. Furthermore, a maximum search algorithm that is immune to local maxima in an FM profile is discussed. Using this FMF and search algorithm, an Automatic focusing system (AFS) tailored for optical tweezers is presented. The system is implemented on personal computers equipped with Pentium 4 processors.
There are several new tools for manipulating microscopic objects. Among them, optical tweezers (OT) has two distinguishing advantages. Firstly, OT can easily release an object without the need of a complicated detaching scheme. Secondly, it is anticipated to manipulate an object with six degrees of freedom. OT is realized by tightly focusing a laser beam on microscopic objects. Grabbing and releasing is easily done by turning a laser beam on and off. For doing a dexterous manipulation on an object, a complicated potential trap must be calculated and applied. We foresee that such calculation method will be developed in the near future. One of the candidates for implementing the calculated trap is scanning optical tweezers (SOT). SOT can be built by using actuators with a scanning frequency in the order of a hundred Hertz. We need fast scanners to stably trap an object. In this study, we present our design of such SOT. The SOT uses piezo-actuated tilt mirror and objective positioner to scan full three-dimensional workspace.
There are increased needs for manipulating microscopic objects. One of enabling technologies is an instrument called optical tweezers (OT) that uses a focused laser beam to trap and move microscopic objects. OT has been shown effective for directly manipulating spherical, cylindrical or axis-symmetrical shapes. For other forms of shapes that do not show any symmetry, there have been works on using micrometer sized balls as a handle to indirectly manipulate the objects. Direct manipulation is difficult because complex trapping potential needs to be calculated to stably trap non-symmetrical shapes. User interfaces for these "indirect" systems use a computer mouse to design a layout of balls for surrounding (holding) an object and a trajectory that describes how these balls as a whole moves. The contained object pushed by these surrounding balls then moves accordingly. In this study, we introduce an intuitive user interface system for manipulating these balls. Using virtual reality gloves, each finger tip position of an operator is used to position control these balls. This user interface system enables the operator to intuitively grasp, move and release irregular formed shapes.