Off-axis digital holographic microscopy (DHM) provides both amplitude and phase images, and so may be used for label-free 3D tracking of micron- and submicron-sized particles of different compositions, including biological cells, strongly absorbing particles, and strongly scattering particles. Contrast is provided by differences in either the real or imaginary parts of the refractive index (phase contrast and absorption) and/or by scattering. While numerous studies have focused on improving resolution in DHM, particularly axial resolution, there has not been studies quantifying the limits of ability to detect and track unresolved particles. This measurement has important implications for microbial detection, including in life-detection missions for space flight. Here we examine the limits of detection of nanosized particles as a function of particle optical properties, microscope optics (including camera well depth and substrate), and data processing techniques.
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