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

Abstract

Single-shot, two-frame, π-shifted spatially multiplexed interference microscopy (π-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 π-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 π radians at the expense of reducing the field of view. Upon subsequent subtractive superimposition of holograms, a π-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, π-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 20 × ∕0.46 NA objective in a regular Olympus BX-60 upright microscope for QPI of different lines of prostate cancer cells and flowing microbeads.

Variational Hilbert Quantitative Phase Imaging

“…In this contribution we are focusing on accurate bi-dimensional single-shot approaches as ability to study full-field dynamic events is fundamental in biomedicine. Single-shot slightly-off axis methods with numerical operators require either constrains on the usable field of view 43 , 47 , object and reference beam intensities 38 , 39 and/or complicated optimization 40 , or are limited to image 1D scanning 49 – 52 . Nonetheless, recently a very elegant approach employing Kramers–Kronig relations was introduced to the QPI and digital holographic imaging in general 53 .…”

“…It attempts the space-bandwidth product optimization by means of full spectral separation of conjugated object lobes, while leaving the autocorrelation term partially overlapped with information carrying cross-correlation terms [38][39][40][41][42][43][44][45][46][47][48][49][50][51][52] . Slightly off-axis phase demodulation is presently facilitated mainly by imposing restrictions on object and reference beams [38][39][40] , utilizing subtraction of two images [41][42][43][44][45][46][47] , employing two wavelengths 48 , or basing on the 1D limited processing [49][50][51][52] , however. In this contribution we are focusing on accurate bi-dimensional single-shot approaches as ability to study full-field dynamic events is fundamental in biomedicine.…”

Single-shot wavelength-multiplexed phase microscopy under Gabor regime in a regular microscope embodiment

“…Building a simple arrangement is of particular significance in DHM since it provides a compact and costeffective solution in a comprehensive and easy-to-use way for QPI. Thus, DHM has been proposed using a single interferometric element such as a thick glass plate 8 , a wedge prism 9 , a Fresnel's biprism 10 , a Lloyd's mirror 11 , a beam splitter cube 12 or a diffraction grating 13 , just to cite some examples. Also following in this uncomplicated and compact way to produce QPI, several attempts have been dedicated to convert-with minimal modifications-a regular white light microscope into a microscope with coherence sensing capabilities.…”

High Bandwidth‐Utilization Digital Holographic Multiplexing: An Approach Using Kramers–Kronig Relations

“…[41] The multiplexing bandwidth utilization in Figure 1f surpasses the current multiplexing limit to 78.5%, which is the highest with the circular pupil function of the diffraction-limited optical imaging system; however, it inevitably leads to the overlap between the AC and CC spectra. It can be reconstructed by acquiring two phase-shifted holograms [50,[54][55][56] but consumes multiframe images or more pixels, reducing at least half of the SBU. The FOV multiplexing is considered as an example in the analysis here.…”