Holotomography: Refractive Index as an Intrinsic Imaging Contrast for 3-D Label-Free Live Cell Imaging

Kim, Doyeon; Lee, Sangyun; Oh, Juntaek; Yang, Su-A; Park, YongKeun

doi:10.1007/978-981-33-6064-8_10

Cited by 27 publications

(26 citation statements)

References 147 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Another interesting possibility is to improve SNR using template-matching techniques based on full spatial and temporal correlations in the 4D spike-induced deformation movie. These could include machine-learning algorithms, along with spatial averaging over the region of interest (ROI) predicted by the 3D cell model from confocal microscopy or holographic tomography (47).…”

Section: Discussionmentioning

confidence: 99%

High-speed interferometric imaging reveals dynamics of neuronal deformation during the action potential

Ling

Boyle

Zuckerman

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Neurons undergo nanometer-scale deformations during action potentials, and the underlying mechanism has been actively debated for decades. Previous observations were limited to a single spot or the cell boundary, while movement across the entire neuron during the action potential remained unclear. Here we report full-field imaging of cellular deformations accompanying the action potential in mammalian neuron somas (−1.8 to 1.4 nm) and neurites (−0.7 to 0.9 nm), using high-speed quantitative phase imaging with a temporal resolution of 0.1 ms and an optical path length sensitivity of <4 pm per pixel. The spike-triggered average, synchronized to electrical recording, demonstrates that the time course of the optical phase changes closely matches the dynamics of the electrical signal. Utilizing the spatial and temporal correlations of the phase signals across the cell, we enhance the detection and segmentation of spiking cells compared to the shot-noise–limited performance of single pixels. Using three-dimensional (3D) cellular morphology extracted via confocal microscopy, we demonstrate that the voltage-dependent changes in the membrane tension induced by ionic repulsion can explain the magnitude, time course, and spatial features of the phase imaging. Our full-field observations of the spike-induced deformations shed light upon the electromechanical coupling mechanism in electrogenic cells and open the door to noninvasive label-free imaging of neural signaling.

show abstract

Section: Discussionmentioning

confidence: 99%

High-speed interferometric imaging reveals dynamics of neuronal deformation during the action potential

Ling

Boyle

Zuckerman

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…We next interrogated the optical characteristics of the RfA2-based structures distributed throughout the bodies of our engineered cells. To achieve this goal, we probed RfA2-expressing cells with a commercially available holotomography microscope (see Methods for full details). ,− The representative phase microscopy images collected for our engineered cells confirmed the presence of variable-sized subcellular RfA2-based structures with a substantial contrast relative to their immediate surroundings, i.e., brighter appearances due to stronger light scattering (Figure A, left, and see also Figure S9). The representative three-dimensional refractive index maps formulated from multiple images of our RfA2-expressing cells revealed average global refractive indices of ∼1.42 ± 0.01, which were comparable to those previously found for RfA1-expressing cells (Figure A, right, and see also Figure S9 and Video S1).…”

Section: Resultsmentioning

confidence: 92%

“…We furthermore interrogated the optical characteristics of the RfA2-based structures found within the EVs released by our engineered cells. To achieve this goal, we collected the vesicles from the cell culture media using established methodologies , and again probed them with a commercially available holotomography microscope (see Methods for full details). ,− The representative phase microscopy images collected for EVs from RfA2-expressing cells confirmed the presence of internal RfA2-based light-scattering structures, which featured a substantial contrast relative to their immediate surroundings (Figure A, left, and see also Figure S11). The representative three-dimensional refractive index maps formulated from multiple images of our RfA2-containing EVs revealed average global refractive indices of ∼1.41 ± 0.01, which were similar to those quantified for the engineered cells (Figure A, right, and see also Figure S11 and Video S2).…”

Section: Resultsmentioning

confidence: 99%

Squid Skin Cell-Inspired Refractive Index Mapping of Cells, Vesicles, and Nanostructures

Chatterjee

Pratakshya

Kwansa

et al. 2023

ACS Biomater. Sci. Eng.

View full text Add to dashboard Cite

The fascination with the optical properties of naturally occurring systems has been driven in part by nature's ability to produce a diverse palette of vibrant colors from a relatively small number of common structural motifs. Within this context, some cephalopod species have evolved skin cells called iridophores and leucophores whose constituent ultrastructures reflect light in different ways but are composed of the same high refractive index material�a protein called reflectin. Although such natural optical systems have attracted much research interest, measuring the refractive indices of biomaterial-based structures across multiple different environments and establishing theoretical frameworks for accurately describing the obtained refractive index values has proven challenging. Herein, we employ a synergistic combination of experimental and computational methodologies to systematically map the three-dimensional refractive index distributions of model self-assembled reflectin-based structures both in vivo and in vitro. When considered together, our findings may improve understanding of squid skin cell functionality, augment existing methods for characterizing protein-based optical materials, and expand the utility of emerging holotomographic microscopy techniques.

show abstract

“…The partial coherence also makes the measurements robust to speckle noise that commonly affects label-free imaging methods. Relative to the interference-based optical designs (7,67,68), our non-interferometric design achieves better robustness to speckle noise by trading-off the sensitivity to large-scale variations in the specimen (23,31) that correspond to low spatial frequencies.…”

Section: Discussionmentioning

confidence: 99%

uPTI: uniaxial permittivity tensor imaging of intrinsic density and anisotropy

Yeh

Ivanov

Guo

et al. 2020

Preprint

View full text Add to dashboard Cite

Architecture of biological systems is important for their homeostatic functions and is predictive of pathology. Volumetric imaging of intrinsic density, anisotropy, and 3D orientation of cell and tissue components is informative, but still challenging. These physical properties can be described succinctly by the volumetric distribution of the specimen’s permittivity tensor (PT). We report uniaxial permittivity tensor imaging (uPTI), a novel approach for label-free volumetric imaging with diffraction-limited resolution. uPTI combines the oblique illumination and the polarization imaging with high numerical apertures to encode the specimen’s permittivity tensor into intensity modulations, which are decoded with a novel vector diffraction model and a multi-channel convex optimization. The uPTI volumes of polystyrene beads and laser-fabricated anisotropic glass targets show that 3D uPT measurements are quantitative, with diffraction-limited spatial resolution of 0.23 × 0.23 × 0.8 μm3. Automated 2D and 3D imaging of a mouse brain section reveals anatomy at multiple scales (cm – 250 nm) and demonstrates that uPTI enables analysis of the density, anisotropy, and orientation of axon bundles and single axons. We multiplex uPTI with fluorescence de-convolution microscopy to enable correlative analysis of physical and molecular architecture of iPSC-derived cardiomyocytes. uPTI can be added to an existing widefield microscope as a low cost module. We share our implementation of the forward model and optimization algorithms via open source repository. Collectively, the reported advances in optical design, image formation, reconstruction algorithms, and biological interpretation open multiple avenues to study architecture of organelles, cells and tissue sections, including human cells and tissues that are challenging to label.

show abstract

Holotomography: Refractive Index as an Intrinsic Imaging Contrast for 3-D Label-Free Live Cell Imaging

Cited by 27 publications

References 147 publications

High-speed interferometric imaging reveals dynamics of neuronal deformation during the action potential

High-speed interferometric imaging reveals dynamics of neuronal deformation during the action potential

Squid Skin Cell-Inspired Refractive Index Mapping of Cells, Vesicles, and Nanostructures

uPTI: uniaxial permittivity tensor imaging of intrinsic density and anisotropy

Contact Info

Product

Resources

About