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Detectability of unresolved particles in off-axis digital holographic microscopy

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Abstract

Off-axis digital holographic microscopy (DHM) provides both amplitude and phase images, and so it may be used for label-free 3D tracking of micro- and nano-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 phase contrast and improving resolution in DHM, particularly axial resolution, absent have been studies quantifying the limits of detection for unresolved particles. This limit 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 and find that DHM provides contrast in both amplitude and phase for unresolved spheres, in rough agreement with Mie theory scattering cross-sections. Amplitude reconstructions are more useful than phase for low-index spheres and should not be neglected in DHM analysis.

© 2024 Optica Publishing Group

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Supplementary Material (1)

NameDescription
Supplement 1       Supplementary Figures S1-S3

Data availability

The raw holograms for the datasets used here are deposited in a public depository at Data Dryad [58]. Other data, such as reconstructions, are available from the authors upon request. The software packages used are all open source and are available at the following sites: acquisition software, DHMx: [59]; Reconstruction Fiji plug-ins: [60].

58. J. Nadeau, “Detectability of unresolved particles in off-axis digital holographic microscopy [dataset],” Dryad, 2024, https://doi.org/10.5061/dryad.9cnp5hqr7.

59. S. F. Fregoso, “DHMx Software Suite,” Github, 2020, https://github.com/dhm-org/dhm_suite.

60. Various repositories, https://github.com/sudgy/.

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Figures (9)

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Equations (11)

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