RESEARCH

Apoptosis

Apoptosis induced by a cytopathic agent (bioactive phospholipids) in rat sarcoma cell observed with Coherence-controlled holographic microscope (obj. 20x/NA 0.4). Pseudocolour quantitative phase images, time in minutes.


Quantitative phase imaging

Label-free non-invasive quantitative phase imaging of K2 rat sarcoma cells proliferation by CCHM. Colour look-up table represents cellular dry-mass density in pg/µm^2. Cell mitosis accompanied by density increase is indicated by red colour in this scale.

                                          


Scattering Layer: Difference in Image Quality with Low and High Coherence of Illumination

Comparison of phase images of MCF-7 cells through turbid 0.1% biologically active phospholipids (BAPs) emulsion. CCHM equipped with (a) low-coherence illumination (halogen lamp), (b) coherent illumination without averaging (Fianium WhiteLase Supercontinuum laser), and (c) coherent illumination with average of 10 frames. The advantage of low-coherence illumination for a single-shot image of cells is clearly demonstrated. Pseudocolor quantitative phase imaging (QPI), objectives 20 × ∕0.5.

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Cell Reactions to Biologically Active Phospholipids

A colony of human colorectal adenocarcinoma cells DLD-1 was followed in the static chamber after exposure to 0.15% BAPs in the culture medium. Three-hour intervals between (a) and (d) disclosed that apparent shrinkage of the whole colony began after 3 h, followed by degradation of some cells not only at the edge of the colony but also inside, with clear condensation of all cell bodies in the colony marking their death. Pseudocolor QPI, calibration bar expresses dry mass density in pg∕μm2, objectives 20 × ∕0.5.

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Cell Cycle Measurement

Left: Cell cycle measurement. Steady gain on the weight of mother (M) and first (D1, D2) and second (D3, D4) generation of daughter cells indicates continuous cell cycling. Right: The boxplot indicates Doubling Time of cells exposed to different nutritional conditions: full medium (FBS 10%) or basic serum-free (FBS 0%). Doubling Time is calculated from the growth rate of each cell for estimation of time that a cell potentially needs for doubling its mass and thus presumably finishing the cell cycle. The graph demonstrates higher growth rate and consequently shorter cell cycle in full medium. Dot markers in boxes indicate mean, centreline is median, top of the box is 25th and bottom 75th percentile line, whiskers indicate 5th and 95th percentiles, outliers are plotted, 47 samples from each culture, significance was tested using unpaired t-test (***: p<0.001). 


Optical setup of Q-Phase

Optical setup of Coherence-controlled holographic microscope alias Q-Phase. Light source (S), relay lens (L), beam splitters (BS), condensers (C), specimen (SP), reference object (RO), objectives (O), tube lenses (TL), diffraction grating (DG), output lenses (OL), output plane (OP), detector (D). 


Image processing

Image processing made by coherence-controlled holographic microscope (CCHM) : (a) hologram of a live cell, (b) the spatial-frequency spectrum with the windowing operation around the carrier frequency, (c) amplitude image, (d) phase image, and (e) quantitative phase image.

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Polarization sensitive phase-shifting Mirau interferometry using a liquid crystal variable retarder

(a) Schema of the polarization adapted phase-shifting (P-S) Mirau interferometer. The P-S is based on a liquid crystal light modulation, imposed on an unaffected reference wave by the Liquid Crystal Variable Retarder (LCVR). (b-e) Experimental testing of the polarization adapted Mirau interferometer. (b) Phase reconstruction of the array of microlenses, (c) 3D illustration of the unwrapped phase, (d) height profiles across the valleys of the microlenses (Sections A and B), (e) height profiles across the tops of the microlenses (Sections C and D).

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Aberration resistant axial localization using a self-imaging of vortices

(a) Experimental setup for aberration resistant axial localization by the rotating Double-Helix Point Spread Function (DH PSF). (b) Rotating imaging of polystyrene beads, 1 micrometer in diameter, suspended in glass capillary tube. (c-e) Rotating images of polystyrene beads in magnified portions of the entire field of view (b). 

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Multimode fiber: Light-sheet microscopy at the tip of a needle

(a) A simplified scheme of the LS delivery using the MMF. The insets (I–VI) show the yz-plane profiles of the GB, BB and SI-BB LS in focus (x = 50 μm from fibre facet; LS scanning shown in Media. 1–3) and out of focus (x = 100 μm from fibre facet; LS scanning shown in Media. 4–6). (b) A strategy for obtaining the SLM hologram generating the GB LS at the fiber output. (c) A procedure converting the GB LS to a BB/SI-BB LS.

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