A single exposure of digital Gabor holography (DGH) is used for simultaneous three-dimensional measurement of
particle position. The particle sample is set up such that its position can be electro-mechanically manipulated using
calibrated piezoelectric transducers in both the lateral and axial directions. The central position of the reconstructed
image of the particle is determined by low-pass filtering, thresholding, and center-of-mass calculation. We have obtained
less than 20 nm resolution in both the lateral and axial directions in a direct and unambiguous manner. The method is
applied to calibration of optical trap strength.
It has recently been demonstrated that diode laser bars can be used to not only optically trap red blood cells in flowing
microfluidic systems but also, stretch, bend, and rotate them. To predict the complex cell behavior at different locations
along a linear trap, 3D optical force characterization is required. The driving force for cells or colloidal particles within
an optical trap is the thermal Brownian force where particle fluctuations can be considered a stochastic process. For
optical force quantification, we combine diode laser bar optical trapping with Gabor digital holography imaging to
perform subpixel resolution measurements of micron-sized particles positions along the laser bar. Here, diffraction
patterns produced by trapped particles illuminated by a He-Ne laser are recorded with a CMOS sensor at 1000 fps where
particle beam position reconstruction is performed using the angular spectrum method and centroid position detection.
3D optical forces are then calculated by three calibration methods: the equipartition theorem, Boltzmann probability
distribution, and power spectral density analysis for each particle in the trap. This simple approach for 3D tracking and
optical control can be implemented on any transmission microscope by adding a laser beam as the illumination source
instead of a white light source.
Structured illumination microscopy (SIM) is a valuable tool for three-dimensional microscopy and has numerous applications in bioscience. Its success has been limited to static objects, though, as three sequential image acquisitions are required per final processed, focused image. To overcome this problem we have developed a multicolored grid which when used in tandem with a color camera is capable of performing SIM with just a single exposure. Images and movies demonstrating optical sectioning of three-dimensional objects are presented, and results of applying color SIM for wide-field focused imaging are compared to those of SIM. From computer modeling and analytical calculations a theoretical estimate of the maximum observable object velocity in both the lateral and axial directions is available, implying that the new method will be capable of imaging a variety of live objects. Sample images of the technique applied to lens paper and a pigeon feather are included to show both advantages and disadvantages of CSIM.