Microscopic Object Classification through Passive Motion Observations with Holographic Microscopy

Abstract

Digital holographic microscopy provides the ability to observe throughout a volume that is large compared to its resolution without the need to actively refocus to capture the entire volume. This enables simultaneous observations of large numbers of small objects within such a volume. We have constructed a microscope that can observe a volume of 0.4 µm × 0.4 µm × 1.0 µm with submicrometer resolution (in xy) and 2 µm resolution (in z) for observation of microorganisms and minerals in liquid environments on Earth and on potential planetary missions. Because environmental samples are likely to contain mixtures of inorganics and microorganisms of comparable sizes near the resolution limit of the instrument, discrimination between living and non-living objects may be difficult. The active 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. We demonstrate the use of passive motion to make this discrimination by evaluating diffusion and buoyancy characteristics of cells, styrene beads, alumina particles, and gas-filled vesicles of micron scale in the field of view.

The Use of Immersion Oil in Parasitology Light Microscopic Examination

“…Light microscopy is a powerful tool for examining micrometric objects (unseen by naked eyes) [1,2], organic (motile or alive) or non-organic (non-motile or in-organic) [3], across a wide range of applications in systems Biology [4] by providing an enlarged image of small structure that deliberately interact with visible light [5], for example their light-absorption, light-reflection and even light-scattering. Light microscopy is used to make small structures and samples visible by using the aid of light, naturally or electrically [1][2][3][4][5]. Contemporary light microscopes are able to magnify objects up to about a thousand times.…”

“…This simple microscopic approach is useful to help observer to (1) get the larger picture of the object for identification, (2) analyze and understand the morphologic appearance of the sample for differentiation and even perhaps (3) to explore the material construction in more detail. Light microscope also allows examiners to observe sophisticated scrutiny of the microscopic world, such as how substances diffuse across a cell membrane [1].…”

Digital holographic microscopy for bacterial species classification and motility characterization

“…Many of these works rely on deep learning the quantitative phase images of cells, which are produced by off-axis DHM or similar techniques and are typically not accessible by inline DHM. There are also reports on machine learning bacteria's motility characteristics to identify bacterial cells, especially in studies aimed at searching for extraterrestrial life 12,13 . However, using motility characteristics to classify species may not be suitable for investigating the interactions of bacteria, as their swimming behavior could be varied in the mixture, different from when they are the sole species in the environment.…”