Microscopic Object Classification through Passive Motion Observations with Holographic Microscopy

Digital holographic microscopy provides the ability to observe throughout a volume that is large compared to its resolution element without the need to refocus through the volume. This capability enables simultaneous observations of large numbers of small objects within such a volume. We have constructed a microscope that can observe a volume 0.4 × 0.4 × 1.0 μm with submicrometer resolution for observation of microorganisms and minerals in liquid environments in earth and on potential planetary missions. Because environmental samples are likely to contain mixtures of inorganics and microorganisms that are of comparable sizes near the resolution limit of the instrument, discrimination of living and non-living objects may be difficult. The motion of motile organisms can be used to readily distinguish them from non-motile objects (live or inorganic), but additional methods are required to distinguish non-motile organisms and inorganic objects that are of comparable size but different composition and structure. In this paper we evaluate the use of passive motion to make this discrimination by evaluating diffusion and buoyancy characteristics of objects in the field of view.