A Simple L ensless Digital Holographic M icroscope (2012)

Abstract: Transparent objects can be imaged with high resolution along the optical axis by numerical reconstruction of digitally recorded holograms. A simple lensless digital holographic microscope is demonstrated to image optically transparent phase objects. © 2011 Optical Society of AmericaOCIS codes: (090.1995) Digital holography; (180.6900) Three-dimensional microscopy; (120.5050) Phase measurement 1. Introduction Optically transparent material such as biological cells can be visualized by digital holographic microscopy that has traditionally been done with phase contrast methods such as Zernike phase contrast, DIC, and other interferometric methods. A major advantage of digital holographic microscopy is recording both the amplitude and phase of the object, simultaneously, leading to numerical focusing and quantitative estimation of the phase. A quasi-Fourier hologram can be described as a hologram recorded with a point source acting as a spherical reference located in the same plane as that of the object and reconstructed later using a plane wave [1]. To record high resolution digital holograms, a CCD (charge-coupled device) sensor with high numerical aperture is needed. The numerical aperture of a CCD sensor can be increased by reducing the object to sensor distance or using a large format sensor. The object to sensor distance is limited by the adjacent pixel distance of the sensor resulting from the requirement to sample the hologram at or above the Nyquist limit. The spatial frequencies of the hologram can be minimized by the use of a Fourier holography setup. The basic principle in the lensless quasi Fourier transform holography is to match the spherical phase factors that are due to the propagation of the object wave and the reference wave at the hologram plane. The use of a point source illumination ensures the full spatial bandwidth of the sampling medium like a CCD [2-4]. For a plane reference wave, for example, the angle between the reference beam and the object beam varies over the sensor surface, therefore, in some places the full spatial bandwidth is not utilized under the assumption of Nyquist sampling theorem. Computation and reconstruction of a digitized hologram has been demonstrated in the literature [5-7] and its application to microscopy and phase imaging [[8-10] are providing a new tool for quantitative estimation of optical properties otherwise hard to measure in certain applications. 2. Principle A simple optical scheme for holographic recording of phase objects is shown in Fig. 1. Two mutually coherent point sources are placed laterally shifted and equidistant from the recording plane, representing two spherically diverging beams. The object to be imaged is placed in one of the spherical beams resulting in a blurred and magnified image of the object at the recording plane. The object to point source distance determines the magnification of the recorded image. Interference pattern of the two beams is recorded on a CCD with a linear carrier spatial frequency given by displacement of the two point sources. A quasi lensless Fourier transform hologram is then sampled by the CCD sensor and further processed numerically for phase imaging. As shown in Fig. 1, a phase object is illuminated by a spherically expanding beam that can be written at the object plane as