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Phase Uncertainty in Digital Holographic Microscopy Measurements in the Presence of Solution Flow Conditions



Alexander Brand


Digital holographic microscopy (DHM) is a surface topography measurement technique with reported sub- nanometer vertical resolution. While made commercially available recently, few studies have evaluated the uncertainty or noise in the phase measurement by the DHM. As current research is using the DHM to monitor surface topography changes of dissolving materials under flowing water conditions, it is necessary to evaluate the effect of water and flow rate on the uncertainty in the measurement, as all previous studies have considered uncertainty in measurements made in air. Uncertainty in this study was concerned with the temporal standard deviation per pixel of the reconstructed phase. Considering the effects of solution flow rate, magnification, objective lens type (air or immersion), and experimental configuration, measurements under static conditions in air and in water with an immersion lens yielded the smallest amount of uncertainty (mean of ≤ 0.5 nm up to 40x). Increasing the water flow rate resulted in an increase in mean uncertainty to ≤ 0.6 nm up to 40x with an immersion lens. Observations through a glass window at 20x of a sample in flowing water also yielded increasing uncertainty, with mean values of ≤ 0.5 nm, ≤ 0.8 nm, and ≤ 1.1 nm for flow rates of 0 mL min-1, 15 mL min-1, and 33 mL min-1. Different hologram acquisition rates (12.5 s-1 and 25 s-1) did not significantly impact the uncertainty in the phase. Collecting holograms in single wavelength versus dual wavelength modes did impact the uncertainty, with the mean uncertainty at 10x for the same wavelength being ≤ 0.5 nm from the single wavelength mode compared to ≤ 1.5 nm from the dual wavelength mode. Applying the quantified uncertainty to simulated dissolution data, lower limits of measured dissolution rates were found below which the measured data may not be distinguishable from the uncertainty in the measurement. The limiting surface normal dissolution velocity is -10-11.7 m s-1 for experiments wi
Journal of Research (NIST JRES) -


digital holographic microscopy, uncertainty, quantitative phase microscopy, dissolution, precipitation


Brand, A. (2017), Phase Uncertainty in Digital Holographic Microscopy Measurements in the Presence of Solution Flow Conditions, Journal of Research (NIST JRES), National Institute of Standards and Technology, Gaithersburg, MD, [online], (Accessed April 18, 2024)
Created March 27, 2017, Updated November 10, 2018