scholarly journals Revealing architectural order with quantitative label-free imaging and deep learning

2019 ◽  
Author(s):  
Syuan-Ming Guo ◽  
Li-Hao Yeh ◽  
Jenny Folkesson ◽  
Ivan Ivanov ◽  
Anitha Priya Krishnan ◽  
...  
Keyword(s):  
Quantitative Imaging ◽  
Three Dimensional ◽  
Kidney Tissue ◽  
Live Cells ◽  
Label Free ◽  
Computationally Efficient ◽  
Human Brain Tissue ◽  
Tissue Slices ◽  
Tissue Sections ◽  
Scalable Analysis

Quantitative imaging of biological architecture with fluorescent labels is not as scalable as genomic or proteomic measurements. Here, we combine quantitative label-free imaging and deep neural networks for scalable analysis of complex structures. We reconstruct quantitative three-dimensional density, anisotropy, and orientation in live cells and tissue slices from polarization- and depth-resolved images. We report a computationally efficient variant of U-Net architecture that predicts a 3D fluorescent structure from its morphology and physical properties. We evaluate the performance of our models by predicting F-actin and nuclei in mouse kidney tissue. Further, we report label-free imaging of axon tracts and predict level of myelination in human brain tissue sections. We demonstrate the model's ability to rescue inconsistent labeling. We anticipate that the proposed approach will enable quantitative analysis of architectural order across scales of organelles to tissues.

scholarly journals Revealing architectural order with quantitative label-free imaging and deep learning

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Syuan-Ming Guo ◽  
Li-Hao Yeh ◽  
Jenny Folkesson ◽  
Ivan E Ivanov ◽  
Anitha P Krishnan ◽  
...  
Keyword(s):  
Spatial Scales ◽  
Brain Regions ◽  
Three Dimensions ◽  
Live Cells ◽  
Label Free ◽  
Computationally Efficient ◽  
Human Brain Tissue ◽  
Tissue Slices ◽  
Efficient Prediction ◽  
Fluorescence Images

We report quantitative label-free imaging with phase and polarization (QLIPP) for simultaneous measurement of density, anisotropy, and orientation of structures in unlabeled live cells and tissue slices. We combine QLIPP with deep neural networks to predict fluorescence images of diverse cell and tissue structures. QLIPP images reveal anatomical regions and axon tract orientation in prenatal human brain tissue sections that are not visible using brightfield imaging. We report a variant of U-Net architecture, multi-channel 2.5D U-Net, for computationally efficient prediction of fluorescence images in three dimensions and over large fields of view. Further, we develop data normalization methods for accurate prediction of myelin distribution over large brain regions. We show that experimental defects in labeling the human tissue can be rescued with quantitative label-free imaging and neural network model. We anticipate that the proposed method will enable new studies of architectural order at spatial scales ranging from organelles to tissue.

Automated 3-D cell population analysis in thick tissue sections using laser-scanning confocal-microscopy data

1993 ◽  
Vol 51 ◽  
pp. 562-563
Author(s):  
Hakan Ancin
Keyword(s):  
Laser Scanning ◽  
Population Analysis ◽  
Three Dimensional ◽  
Large Data ◽  
Laser Scanning Confocal Microscopy ◽  
Tissue Slices ◽  
Scanning Confocal Microscopy ◽  
Tissue Sections ◽  
Spatially Adaptive ◽  
Microscopy Data

This paper presents methods for performing detailed quantitative automated three dimensional (3-D) analysis of cell populations in thick tissue sections while preserving the relative 3-D locations of cells. Specifically, the method disambiguates overlapping clusters of cells, and accurately measures the volume, 3-D location, and shape parameters for each cell. Finally, the entire population of cells is analyzed to detect patterns and groupings with respect to various combinations of cell properties. All of the above is accomplished with zero subjective bias.In this method, a laser-scanning confocal light microscope (LSCM) is used to collect optical sections through the entire thickness (100 - 500μm) of fluorescently-labelled tissue slices. The acquired stack of optical slices is first subjected to axial deblurring using the expectation maximization (EM) algorithm. The resulting isotropic 3-D image is segmented using a spatially-adaptive Poisson based image segmentation algorithm with region-dependent smoothing parameters. Extracting the voxels that were labelled as "foreground" into an active voxel data structure results in a large data reduction.

scholarly journals Mitotic Chromosomes in Live Cells Characterized Using High-Speed and Label-Free Optical Diffraction Tomography

Cells ◽  
2019 ◽  
Vol 8 (11) ◽  
pp. 1368 ◽  
Author(s):  
Kim ◽  
Lee ◽  
Fujii ◽  
Lee ◽  
Lee ◽  
...  
Keyword(s):  
Diffusion Coefficient ◽  
Three Dimensional ◽  
Laser Scanning Microscopy ◽  
Optical Diffraction ◽  
Live Cells ◽  
Confocal Laser ◽  
Diffraction Tomography ◽  
Label Free ◽  
Mitotic Chromosomes ◽  
Optical Diffraction Tomography

The cell nucleus is a three-dimensional, dynamic organelle organized into subnuclear compartments such as chromatin and nucleoli. The structure and function of these compartments are maintained by diffusion and interactions between related factors as well as by dynamic and structural changes. Recent studies using fluorescent microscopic techniques suggest that protein factors can access and are freely mobile in heterochromatin and in mitotic chromosomes, despite their densely packed structure. However, the physicochemical properties of the chromosome during cell division are not fully understood. In the present study, characteristic properties such as the refractive index (RI), volume of the mitotic chromosomes, and diffusion coefficient (D) of fluorescent probes inside the chromosome were quantified using an approach combining label-free optical diffraction tomography with complementary confocal laser-scanning microscopy and fluorescence correlation spectroscopy. Variations in these parameters correlated with osmotic conditions, suggesting that changes in RI are consistent with those of the diffusion coefficient for mitotic chromosomes and cytosol. Serial RI tomography images of chromosomes in live cells during mitosis were compared with three-dimensional confocal micrographs to demonstrate that compaction and decompaction of chromosomes induced by osmotic change were characterized by linked changes in chromosome RI, volume, and the mobilities of fluorescent proteins.

scholarly journals In vivo, label-free, three-dimensional quantitative imaging of kidney microcirculation using Doppler optical coherence tomography

2011 ◽  
Vol 91 (11) ◽  
pp. 1596-1604 ◽  
Author(s):  
Jeremiah Wierwille ◽  
Peter M Andrews ◽  
Maristela L Onozato ◽  
James Jiang ◽  
Alex Cable ◽  
...  
Keyword(s):  
Optical Coherence Tomography ◽  
Quantitative Imaging ◽  
Three Dimensional ◽  
Optical Coherence ◽  
Label Free ◽  
Doppler Optical Coherence Tomography

scholarly journals Physicochemical Properties of Chromosomes in Live Cells Characterized by Label-Free Imaging and Fluorescence Correlation Spectroscopy

2019 ◽  
Author(s):  
Tae-Keun Kim ◽  
Byong-Wook Lee ◽  
Fumihiko Fujii ◽  
Kee-Hang Lee ◽  
YongKeun Park ◽  
...  
Keyword(s):  
Physicochemical Properties ◽  
Fluorescence Correlation Spectroscopy ◽  
Three Dimensional ◽  
Correlation Spectroscopy ◽  
Laser Scanning Microscopy ◽  
Optical Diffraction ◽  
Live Cells ◽  
Label Free ◽  
Mitotic Chromosomes ◽  
Fluorescence Correlation

AbstractThe cell nucleus is a three-dimensional, dynamic organelle that is organized into many subnuclear bodies, such as chromatin and nucleoli. The structure and function of these bodies is maintained by diffusion and interactions between related factors as well as dynamic and structural changes. Recent studies using fluorescent microscopic techniques suggest that protein factors can access and are freely mobile in mitotic chromosomes, despite their densely packed structure. However, the physicochemical properties of the chromosome itself during cell division are not yet fully understood. Physical parameters, such as the refractive index (RI), volume of the mitotic chromosome, and diffusion coefficients of fluorescent probes inside the chromosome were quantified using an approach combining label-free optical diffraction tomography with complementary confocal laser scanning microscopy and fluorescence correlation spectroscopy. Variance in these parameters correlated among various osmotic conditions, suggesting that changes in RI are consistent with those in the diffusion coefficient for mitotic chromosomes and cytosol. Serial RI tomography images of chromosomes in live cells during mitosis were compared with three-dimensional confocal micrographs to demonstrate that compaction and decompaction of chromosomes induced by osmotic change were characterized by linked changes in chromosome RI, volume, and the mobility of fluorescent proteins.

scholarly journals Live Cell Partial Wave Spectroscopic microscopy: Label-free Imaging of the Native, Living Cellular Nano-architecture

2016 ◽  
Author(s):  
L. M. Almassalha ◽  
G. M. Bauer ◽  
J. Chandler ◽  
S. Gladstein ◽  
L. Cherkezya ◽  
...  
Keyword(s):  
Partial Wave ◽  
Chromatin Structure ◽  
Imaging Technique ◽  
Quantitative Imaging ◽  
Higher Order ◽  
Live Cell ◽  
Live Cells ◽  
Molecular Function ◽  
Metabolic Function ◽  
Label Free

AbstractThe organization of chromatin is a regulator of molecular processes including transcription, replication, and DNA repair. The structures within chromatin that regulate these processes span from the nucleosomal (10nm) to the chromosomal (>200nm) levels, with little known about the dynamics of chromatin structure between these scales due to a lack of quantitative imaging technique in live cells. Previous work using Partial Wave Spectroscopic (PWS) microscopy, a quantitative imaging technique with sensitivity to macromolecular organization between 20-200nm, has shown that transformation of chromatin at these length scales is a fundamental event during carcinogenesis. As the dynamics of chromatin likely play a critical regulatory role in cellular function, it is critical to develop live-cell imaging techniques that can probe the real-time temporal behavior of the chromatin nano-architecture. Therefore, we developed a live cell PWS technique which allows high-throughput, label-free study of the causal relationship between nanoscale organization and molecular function in real-time. In this work, we employ live cell PWS to study the change in chromatin structure due to DNA damage and expand on the link between metabolic function and the structure of higher-order chromatin. In particular, we studied the temporal changes to chromatin during UV light exposure, show that live cell DNA binding dyes induce damage to chromatin within seconds, and demonstrate a direct link between higher-order chromatin structure and mitochondrial membrane potential. Since biological function is tightly paired with structure, live cell PWS is a powerful tool to study the nanoscale structure-function relationship in live cells.Significance StatementChromatin is one of the most critical structures within the cell because it houses most genetic information. Its structure is well understood at the nucleosomal (<20nm) and chromosomal (>200nm) levels, however, due to the lack of quantitative imaging modalities to study this organization, little is known about the higher-order structure between these length scales in live cells. We present a label-free technique, live cell Partial Wave Spectroscopic (PWS) microscopy with sensitivity to structures between 20-200nm that can quantify the nano-architecture in live cells. With this technique, we can detect DNA fragmentation and expand on the link between metabolic function and higher-order chromatin structure. Live cell PWS allows high-throughput, label-free study of the causal relationship between nanoscale organization and molecular function in live cells.

scholarly journals Correlative three-dimensional fluorescence and refractive index tomography: bridging the gap between molecular specificity and quantitative bioimaging

2017 ◽  
Author(s):  
Kyoohyun Kim ◽  
Wei Sun Park ◽  
Sangchan Na ◽  
Sangbum Kim ◽  
Taehong Kim ◽  
...  
Keyword(s):  
Refractive Index ◽  
Three Dimensional ◽  
Optical Diffraction ◽  
Live Cells ◽  
Specific Information ◽  
Label Free ◽  
Optical Diffraction Tomography ◽  
Molecular Specificity ◽  
3D Fluorescence Microscopy ◽  
Nih 3T3

AbstractOptical diffraction tomography (ODT) provides label-free three-dimensional (3D) refractive index (RI) measurement of biological samples. However, due to the nature of the RI values of biological specimens, ODT has limited access to molecular specific information. Here, we present an optical setup combining ODT with three-channel 3D fluorescence microscopy, to enhance the molecular specificity of the 3D RI measurement. The 3D RI distribution and 3D deconvoluted fluorescence images of HeLa cells and NIH-3T3 cells are measured, and the cross-correlative analysis between RI and fluorescence of live cells are presented.

scholarly journals Label-free high-resolution 3-D imaging of gold nanoparticles inside live cells using optical diffraction tomography

2016 ◽  
Author(s):  
Doyeon Kim ◽  
Nuri Oh ◽  
Kyoohyun Kim ◽  
SangYun Lee ◽  
Chan-Gi Pack ◽  
...  
Keyword(s):  
Gold Nanoparticles ◽  
Refractive Index ◽  
Quantitative Imaging ◽  
Extended Period ◽  
Optical Diffraction ◽  
Live Cells ◽  
Diffraction Tomography ◽  
Label Free ◽  
Optical Diffraction Tomography ◽  
4T1 Cells

AbstractDelivery of gold nanoparticles (GNPs) into live cells has high potentials, ranging from molecular-specific imaging, photodiagnostics, to photothermal therapy. However, studying the long-term dynamics of cells with GNPs using conventional fluorescence techniques suffers from phototoxicity and photobleaching. Here, we present a method for 3-D imaging of GNPs inside live cells exploiting refractive index (RI) as imaging contrast. Employing optical diffraction tomography, 3-D RI tomograms of live cells with GNPs are precisely measured for an extended period with sub-micrometer resolution. The locations and contents of GNPs in live cells are precisely addressed and quantified due to their distinctly high RI values, which was validated by confocal fluorescence imaging of fluorescent dye conjugated GNPs. In addition, we perform quantitative imaging analysis including the segmentations of GNPs in the cytosol, the volume distributions of aggregated GNPs, and the temporal evolution of GNPs contents in HeLa and 4T1 cells.AbbreviationsGNPsgold nanoparticlesRIrefractive indexODToptical diffraction tomographyDMDdigital micromirror device

scholarly journals Label-free three-dimensional analyses of live cells with deep-learning-based segmentation exploiting refractive index distributions

2021 ◽  
Author(s):  
Jinho Choi ◽  
Hye-Jin Kim ◽  
Gyuhyeon Sim ◽  
Sumin Lee ◽  
Wei Sun Park ◽  
...  
Keyword(s):  
Cell Biology ◽  
Lipid Droplets ◽  
Three Dimensional ◽  
Time Lapse ◽  
Cell Types ◽  
Live Cells ◽  
Label Free ◽  
Subcellular Organelles ◽  
Long Time ◽  
Three Dimensional Segmentation

Visualisations and analyses of cellular and subcellular organelles in biological cells is crucial for the study of cell biology. However, existing imaging methods require the use of exogenous labelling agents, which prevents the long-time assessments of live cells in their native states. Here we propose and experimentally demonstrate three-dimensional segmentation of subcellular organelles in unlabelled live cells, exploiting a 3D U-Net-based architecture. We present the high-precision three-dimensional segmentation of cell membrane, nucleus membrane, nucleoli, and lipid droplets of various cell types. Time-lapse analyses of dynamics of activated immune cells are also analysed using label-free segmentation.

scholarly journals Dissecting lipid droplet biology with coherent Raman scattering microscopy

2021 ◽  
Vol 135 (5) ◽  
Author(s):  
Tao Chen ◽  
Ahmet Yavuz ◽  
Meng C. Wang
Keyword(s):  
Raman Scattering ◽  
Lipid Droplets ◽  
Quantitative Imaging ◽  
Live Cells ◽  
Label Free ◽  
Genetic Screens ◽  
Spatiotemporal Resolution ◽  
Coherent Raman Scattering ◽  
Coherent Raman

ABSTRACT Lipid droplets (LDs) are lipid-rich organelles universally found in most cells. They serve as a key energy reservoir, actively participate in signal transduction and dynamically communicate with other organelles. LD dysfunction has been associated with a variety of diseases. The content level, composition and mobility of LDs are crucial for their physiological and pathological functions, and these different parameters of LDs are subject to regulation by genetic factors and environmental inputs. Coherent Raman scattering (CRS) microscopy utilizes optical nonlinear processes to probe the intrinsic chemical bond vibration, offering label-free, quantitative imaging of lipids in vivo with high chemical specificity and spatiotemporal resolution. In this Review, we provide an overview over the principle of CRS microscopy and its application in tracking different parameters of LDs in live cells and organisms. We also discuss the use of CRS microscopy in genetic screens to discover lipid regulatory mechanisms and in understanding disease-related lipid pathology.

Sign in / Sign up

Export Citation Format

Share Document