Flipping Interferometric Module for Simultaneous Dual-Wavelength Unwrapping of Quantitative Phase Maps of Biological Cells

Abstract

We present an imaging platform for stain-free quantitative imaging of biological cells using a simultaneous dual-wavelength holographic module. We use this module to experimentally solve the problem of 2π phase ambiguities that occurs in spatial locations where the optical thickness of the sample is larger than the wavelength. Thus, the process does not require using digital phase unwrapping that is computational heavy and impairs real-time processing. The proposed method is not limited to sequential acquisition of two quantitative phase maps in different times, but rather allows optical multiplexing of two off-axis holograms on the camera at once, enabling acquisition of fast dynamic processes. The module is simple and portable, making it attractive for clinical use. We demonstrate using the module for quantitative phase imaging of cancer and sperm cells.

CatSper and its CaM‐like Ca 2+ sensor EFCAB9 are necessary for the path chirality of sperm

“…This, in turn, leads to an increase in swimming trajectories in all three dimensions and sperm velocity, resulting in a stronger swimming force, essential for penetration of the zona pellucida. 13 In the past couple of years, several studies used DHM to track sperm cell trajectories in three dimensions 7,34,51,55,[76][77][78][79] to analyze sperm head morphology 56,80,81 or flagellar movement of free-swimming 7,34,76,80 or tethered sperm. 7,82 Most of the used setups differ in the time resolution.…”

Multi-wavelength off-axis digital holographic microscopy with broadly tunable low-coherent sources: theory, performance and limitations

“…To obtain the intensity modulations S m (r) and OPL differences d m (r) introduced by the sample, the other terms of B m (r) (such as R m (r), see equation (4b)) should be eliminated by a preliminary calibration, e.g. collecting a hologram of a flat featureless area on the sample [2,7,29,41]. This calibration also allows to compensate for the phase aberrations of the optical system.…”

“…Spectral bandwidth of a light source is inversely proportional to its coherence length, which affects the size of high-contrast area of the interference pattern and hence the effective field of view (FOV) in off-axis digital holography. MDHM setups have used various types of light sources, including several lasers or diodes coupled with beamsplitters and dichroic mirrors [3,4,10,16,18,[26][27][28][29], a tunable [20] or multi-wavelength laser [9,30], and a broadband light source followed by interchangeable bandpass filters [8] or a tunable filter. The tunable spectral filter may be based on a prism or a diffraction grating in combination with a galvanometric scanning mirror [13][14][15] or a spatial light modulator (SLM) [7,12], or it can be an acousto-optic tunable filter (AOTF) [5,11,17,[21][22][23][24].…”

“…Although the common-path interferometers are more compact and resistant to vibrations and air fluctuations, they are affected by the spatial coherence of the light illuminating the sample and the size of the pinhole used to filter the reference beam, resulting in phase distortions and reduced light intensity [43]. Another common-path technique does not use a pinhole, but requires significant part of the FOV to be empty [27,29]. Thus, there is always a trade-off between temporal and spatial resolution and stability, FOV and multiwavelength acquisition capabilities.…”