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24 August 2019 Angularly resolved, finely sampled elastic scattering measurements of single cells: requirements for robust organelle size extractions
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Abstract

Angularly resolved elastic light scattering is an established technique for probing the average size of organelles in biological tissue and cellular ensembles. Focusing of the incident light to illuminate no more than one cell at a time restricts the minimum forward-scattering angle θmin that can be detected. Series of simulated single-cell angular-scattering patterns have been generated to explore how size estimates vary as a function of θmin. At a setting of θmin  =  20  deg, the size estimates hop unstably between multiple minima in the solution space as simulated noise (mimicking experimentally observed levels) is varied. As θmin is reduced from 20 deg to 10 deg, the instability vanishes, and the variance of estimates near the correct answer also decreases. The simulations thus suggest that robust Mie theory fits to single-cell scattering at 785 nm excitation require measurements down to at least 15 deg. Notably, no such instability was observed at θmin  =  20  deg for narrow bead distributions. Accurate sizing of traditional calibration beads is, therefore, insufficient proof that an angular-scattering system is capable of robust analysis of single cells. Experimental support for the simulation results is also presented using measurements on cells fixed with formaldehyde.

CC BY: © The Authors. Published by SPIE under a Creative Commons Attribution 4.0 Unported License. Distribution or reproduction of this work in whole or in part requires full attribution of the original publication, including its DOI.
Ashley E. Cannaday, Janet Sorrells, and Andrew J. Berger "Angularly resolved, finely sampled elastic scattering measurements of single cells: requirements for robust organelle size extractions," Journal of Biomedical Optics 24(8), 086502 (24 August 2019). https://doi.org/10.1117/1.JBO.24.8.086502
Received: 12 March 2019; Accepted: 24 July 2019; Published: 24 August 2019
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Cited by 4 scholarly publications.
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KEYWORDS
Scattering

Mie scattering

Light scattering

Speckle

Laser scattering

Scatter measurement

Data modeling

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