Single-shot two-frame π-shifted spatially multiplexed interference phase microscopy.

Maciej Trusiak,Piotr Zdankowski,Jose-Angel Picazo-Bueno,Vicente Mico,Krzysztof Patorski

doi:10.1117/1.jbo.24.9.096004

Maciej Trusiak, Piotr Zdankowski + Show 3 more

Open Access

https://doi.org/10.1117/1.jbo.24.9.096004

Copy DOI

Abstract

.Single-shot, two-frame, -shifted spatially multiplexed interference microscopy (-SMIM) is presented as an improvement to previous SMIM implementations, introducing a versatile, robust, fast, and accurate method for cumbersome, noisy, and low-contrast phase object analysis. The proposed -SMIM equips a commercially available nonholographic microscope with a high-speed (video frame rate) enhanced quantitative phase imaging (QPI) capability by properly placing a beam-splitter in the microscope embodiment to simultaneously (in a single shot) record two holograms mutually phase shifted by radians at the expense of reducing the field of view. Upon subsequent subtractive superimposition of holograms, a -hologram is generated with reduced background and improved modulation of interference fringes. These features determine superior phase retrieval quality, obtained by employing the Hilbert spiral transform on the π-hologram, as compared with a single low-quality (low signal-to-noise ratio) hologram analysis. In addition, -SMIM enables accurate in-vivo analysis of high dynamic range phase objects, otherwise measurable only in static regime using time-consuming phase-shifting. The technique has been validated utilizing a NA objective in a regular Olympus BX-60 upright microscope for QPI of different lines of prostate cancer cells and flowing microbeads.

Highlights

Among a suite of modern microscopy techniques, quantitative phase imaging (QPI)[1,2,3] stands out as a vividly blossoming and extremely capable label-free approach
We propose a single-shot two-frame π-shifted spatially multiplexed interference microscopy (π-spatially multiplexed interferometric microscopy (SMIM)) to overcome the limitations imposed by highly scattering samples resulting in low signal-to-noise ratio (SNR) of recorded holograms
We propose to decompose the π-hologram using the enhanced fast empirical mode decomposition (EFEMD)[47] like it is conducted in the first part of the H2PM for single hologram phase analysis.[29,30]

Summary

Introduction

Among a suite of modern microscopy techniques, quantitative phase imaging (QPI)[1,2,3] stands out as a vividly blossoming and extremely capable label-free approach. Application-oriented QPI research provided recently outstanding solutions in numerous exciting biomedical fields, i.e., in neuroscience, methods such as digital holographic microscopy (DHM),[9] spatial light interference microscopy (SLIM),[10] and optical diffraction tomography (ODT);[11] in cell/tissue biology, ODT,[11] SLIM,[10] transport of intensity,[12] Fourier ptychography,[13] and quadriwave interferometry;[14] and in cancer diagnosis, DHM15 and diffraction phase microscopy,[16,17] to name only some approaches. It is worth showcasing the importance of in-flow quantitative phase measurements.[18,19]

Results

Discussion

Conclusion