Object Plane Detection and Phase Retrieval from Single-Shot Holograms using Multi-Wavelength In-Line Holograph

Phase retrieval and the twin-image problem in digital in-line holographic microscopy can be resolved by iterative reconstruction routines. However, recovering the phase properties of an object in a hologram needs an object plane to be chosen correctly for reconstruction. In this work, we present a novel multi-wavelength Gerchberg-Saxton algorithm to determine the object plane using single-shot holograms recorded with multiple wavelengths in an in-line holographic microscope. For micro-sized objects, we verify the object positioning capabilities of the method for various shapes and derive the phase information using synthetic and experimental data. Experimentally, we built a compact digital in-line holographic microscopy setup around a standard optical microscope with a regular RGB-CCD camera and acquire holograms of micro-spheres, E. coli and red blood cells, that are illuminated using three lasers operating at 491 nm, 532 nm and 633 nm, respectively. We demonstrate that our method provides accurate object plane detection and phase retrieval under noisy conditions, e.g., using low-contrast holograms without background normalization. This method allows for automatic positioning and phase retrieval suitable for holographic particle velocimetry, and object tracking in biophysical or colloidal research.