scholarly journals Multiscale label-free volumetric holographic histopathology of thick-tissue slides with subcellular resolution

2020 ◽  
Author(s):  
Herve Hugonnet ◽  
Yeon Wook Kim ◽  
Moosung Lee ◽  
Seungwoo Shin ◽  
Ralph H. Hruban ◽  
...  
Keyword(s):  
Refractive Index ◽  
Present Method ◽  
Imaging Techniques ◽  
Numerical Algorithms ◽  
Biological Tissues ◽  
Phase Imaging ◽  
Label Free ◽  
Volumetric Imaging ◽  
Optical Modes ◽  
Volumetric Reconstruction

ABSTRACTHistopathology relies upon the staining and sectioning of biological tissues, which can be laborious and may cause artefacts and distort tissues. Here, we demonstrate label-free volumetric imaging of thick-tissue slides, exploiting refractive index distributions as intrinsic imaging contrast. The present method systematically exploits label-free quantitative phase imaging techniques, volumetric reconstruction of intrinsic refractive index distributions in tissues, and numerical algorithms for the seamless stitching of multiple 3D tomograms and for reducing scattering-induced image distortion. We demonstrate demonstrated label-free volumetric imaging of thick tissues with the field of view of 2 mm × 1.75 mm × 0.2 mm with a spatial resolution of 170 nm × 170 nm × 1200 nm. The number of optical modes, calculated as the reconstructed volume divided by the size of the point spread function, was approximately 20 Giga voxels. We have also demonstrated that different tumour types, and a variety of precursor lesions and pathologies can be visualized with the present method.Abstract Figure

scholarly journals Holotomography: refractive index as an intrinsic imaging contrast for 3-D label-free live cell imaging

2017 ◽  
Author(s):  
Doyeon Kim ◽  
SangYun Lee ◽  
Moosung Lee ◽  
JunTaek Oh ◽  
Su-A Yang ◽  
...  
Keyword(s):  
Refractive Index ◽  
Cell Biology ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Imaging Techniques ◽  
Live Cell ◽  
Label Free ◽  
Essential Information ◽  
Research Fields ◽  
Intrinsic Imaging

AbstractLive cell imaging provides essential information in the investigation of cell biology and related pathophysiology. Refractive index (RI) can serve as intrinsic optical imaging contrast for 3-D label-free and quantitative live cell imaging, and provide invaluable information to understand various dynamics of cells and tissues for the study of numerous fields. Recently significant advances have been made in imaging methods and analysis approaches utilizing RI, which are now being transferred to biological and medical research fields, providing novel approaches to investigate the pathophysiology of cells. To provide insight how RI can be used as an imaging contrast for imaging of biological specimens, here we provide the basic principle of RI-based imaging techniques and summarize recent progress on applications, ranging from microbiology, hematology, infectious diseases, hematology, and histopathology.

scholarly journals Visualization and label-free quantification of microfluidic mixing using quantitative phase imaging

2017 ◽  
Author(s):  
GwangSik Park ◽  
Dongsik Han ◽  
GwangSu Kim ◽  
Seungwoo Shin ◽  
Kyoohyun Kim ◽  
...  
Keyword(s):  
Chemical Engineering ◽  
Imaging Techniques ◽  
Quantitative Imaging ◽  
Quantitative Information ◽  
Microfluidic Channels ◽  
Phase Imaging ◽  
Label Free ◽  
Quantitative Phase Imaging ◽  
Microfluidic Mixing ◽  
Quantitative Phase

Microfluidic mixing plays a key role in various fields, including biomedicine and chemical engineering. To date, although various approaches for imaging microfluidic mixing have been proposed, they provide only quantitative imaging capability and require for exogenous labeling agents. Quantitative phase imaging techniques, however, circumvent these problems and offer label-free quantitative information about concentration maps of microfluidic mixing. We present the quantitative phase imaging of microfluidic mixing in various types of PDMS microfluidic channels with different geometries; the feasibility of the present method was validated by comparing it with the results obtained by theoretical calculation based on Fick’s law.

scholarly journals Topography and refractometry of sperm cells using SLIM

2017 ◽  
Author(s):  
Lina Liu ◽  
Mikhail E. Kandel ◽  
Marcello Rubessa ◽  
Sierra Schreiber ◽  
Mathew Wheeler ◽  
...  
Keyword(s):  
Refractive Index ◽  
Interference Microscopy ◽  
Phase Imaging ◽  
Label Free ◽  
Quantitative Phase Imaging ◽  
Ensemble Averaging ◽  
Thickness Profile ◽  
Light Interference ◽  
Phase Sensitive

AbstractCharacterization of spermatozoon viability is a common test in treating infertility. Recently, it has been shown that label-free, phase-sensitive imaging can provide a valuable alternative for this type of assay. Here, we employ spatial light interference microscopy (SLIM) to decouple the thickness and refractive index information of individual cells. This procedure was enabled by quantitative phase imaging cells on media of two different refractive indices and using a numerical tool to remove the curvature from the cell tails. This way, we achieved ensemble averaging of topography and refractometry of 100 cells in each of the two groups. The results show that the thickness profile of the cell tail goes down to 150 nm and the refractive index can reach values of 1.6 close to the head.

scholarly journals Multimodal nonlinear microscopy: A powerful label-free method for supporting standard diagnostics on biological tissues

2014 ◽  
Vol 07 (05) ◽  
pp. 1330008 ◽  
Author(s):  
Riccardo Cicchi ◽  
Francesco Saverio Pavone
Keyword(s):  
Laser Scanning ◽  
Imaging Techniques ◽  
Biological Tissues ◽  
Laser Scanning Microscopy ◽  
Tissue Imaging ◽  
Nonlinear Microscopy ◽  
Label Free ◽  
Free Alternative ◽  
Functional Scheme ◽  
Nonlinear Imaging

The large use of nonlinear laser scanning microscopy in the past decade paved the way for potential clinical application of this imaging technique. Modern nonlinear microscopy techniques offer promising label-free solutions to improve diagnostic performances on tissues. In particular, the combination of multiple nonlinear imaging techniques in the same microscope allows integrating morphological with functional information in a morpho-functional scheme. Such approach provides a high-resolution label-free alternative to both histological and immunohistochemical examination of tissues and is becoming increasingly popular among the clinical community. Nevertheless, several technical improvements, including automatic scanning and image analysis, are required before the technique represents a standard diagnostic method. In this review paper, we highlight the capabilities of multimodal nonlinear microscopy for tissue imaging, by providing various examples on colon, arterial and skin tissues. The comparison between images acquired using multimodal nonlinear microscopy and histology shows a good agreement between the two methods. The results demonstrate that multimodal nonlinear microscopy is a powerful label-free alternative to standard histopathological methods and has the potential to find a stable place in the clinical setting in the near future.

scholarly journals Bacteria Single-Cell and Photosensitizer Interaction Revealed by Quantitative Phase Imaging

2021 ◽  
Vol 22 (10) ◽  
pp. 5068
Author(s):  
Igor Buzalewicz ◽  
Agnieszka Ulatowska-Jarża ◽  
Aleksandra Kaczorowska ◽  
Marlena Gąsior-Głogowska ◽  
Halina Podbielska ◽  
...  
Keyword(s):  
Imaging Techniques ◽  
Single Cells ◽  
Three Dimensional ◽  
Bacterial Biofilm ◽  
Phase Imaging ◽  
Label Free ◽  
Biophysical Processes ◽  
Contemporary Medicine

Quantifying changes in bacteria cells in the presence of antibacterial treatment is one of the main challenges facing contemporary medicine; it is a challenge that is relevant for tackling issues pertaining to bacterial biofilm formation that substantially decreases susceptibility to biocidal agents. Three-dimensional label-free imaging and quantitative analysis of bacteria–photosensitizer interactions, crucial for antimicrobial photodynamic therapy, is still limited due to the use of conventional imaging techniques. We present a new method for investigating the alterations in living cells and quantitatively analyzing the process of bacteria photodynamic inactivation. Digital holographic tomography (DHT) was used for in situ examination of the response of Escherichia coli and Staphylococcus aureus to the accumulation of the photosensitizers immobilized in the copolymer revealed by the changes in the 3D refractive index distributions of single cells. Obtained results were confirmed by confocal microscopy and statistical analysis. We demonstrated that DHT enables real-time characterization of the subcellular structures, the biophysical processes, and the induced local changes of the intracellular density in a label-free manner and at sub-micrometer spatial resolution.

scholarly journals Rapid and label-free identification of individual bacterial pathogens exploiting three-dimensional quantitative phase imaging and deep learning

2019 ◽  
Author(s):  
Geon Kim ◽  
Daewoong Ahn ◽  
Minhee Kang ◽  
YoungJu Jo ◽  
Donghun Ryu ◽  
...  
Keyword(s):  
Refractive Index ◽  
Deep Learning ◽  
Infectious Diseases ◽  
Bacterial Pathogens ◽  
Bacterial Species ◽  
Three Dimensional ◽  
Phase Imaging ◽  
Label Free ◽  
Quantitative Phase Imaging ◽  
Quantitative Phase

ABSTRACTFor appropriate treatments of infectious diseases, rapid identification of the pathogens is crucial. Here, we developed a rapid and label-free method for identifying common bacterial pathogens as individual bacteria by using three-dimensional quantitative phase imaging and deep learning. We achieved 95% accuracy in classifying 19 bacterial species by exploiting the rich information in three-dimensional refractive index tomograms with a convolutional neural network classifier. Extensive analysis of the features extracted by the trained classifier was carried out, which supported that our classifier is capable of learning species-dependent characteristics. We also confirmed that utilizing three-dimensional refractive index tomograms was crucial for identification ability compared to two-dimensional imaging. This method, which does not require time-consuming culture, shows high feasibility for diagnosing patients with infectious diseases who would benefit from immediate and adequate antibiotic treatment.

scholarly journals High-speed multifocus phase imaging in thick tissue

2021 ◽  
Author(s):  
Sheng Xiao ◽  
Shuqi Zheng ◽  
Jerome Mertz
Keyword(s):  
High Speed ◽  
Imaging Techniques ◽  
Index Of Refraction ◽  
Phase Imaging ◽  
Biological Processes ◽  
Thick Sample ◽  
Volumetric Imaging ◽  
Phase Microscopy ◽  
Large Numbers ◽  
Qualitative Phase

Phase microscopy is widely used to image unstained biological samples. However, most phase imaging techniques require transmission geometries, making them unsuited for thick sample applications. Moreover, when applied to volumetric imaging, phase imaging generally requires large numbers of measurements, often making it too slow to capture live biological processes with fast 3D index-of-refraction variations. By combining oblique back-illumination microscopy and a z-splitter prism, we perform phase imaging that is both epi-mode and multifocus, enabling high-speed 3D phase imaging in thick, scattering tissues with a single camera. We demonstrate here 3D qualitative phase imaging of blood flow in chick embryos over a field of view of 546 × 546 × 137 μm3 at speeds up to 47 Hz.

scholarly journals Epi-illumination gradient light interference microscopy for imaging opaque structures

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Mikhail E. Kandel ◽  
Chenfei Hu ◽  
Ghazal Naseri Kouzehgarani ◽  
Eunjung Min ◽  
Kathryn Michele Sullivan ◽  
...  
Keyword(s):  
Multiple Scattering ◽  
Imaging Modality ◽  
Single Cells ◽  
Scattering Problem ◽  
Biological Tissues ◽  
Interference Microscopy ◽  
Model Organisms ◽  
Phase Imaging ◽  
Label Free ◽  
Light Interference

Abstract Multiple scattering and absorption limit the depth at which biological tissues can be imaged with light. In thick unlabeled specimens, multiple scattering randomizes the phase of the field and absorption attenuates light that travels long optical paths. These obstacles limit the performance of transmission imaging. To mitigate these challenges, we developed an epi-illumination gradient light interference microscope (epi-GLIM) as a label-free phase imaging modality applicable to bulk or opaque samples. Epi-GLIM enables studying turbid structures that are hundreds of microns thick and otherwise opaque to transmitted light. We demonstrate this approach with a variety of man-made and biological samples that are incompatible with imaging in a transmission geometry: semiconductors wafers, specimens on opaque and birefringent substrates, cells in microplates, and bulk tissues. We demonstrate that the epi-GLIM data can be used to solve the inverse scattering problem and reconstruct the tomography of single cells and model organisms.

scholarly journals Melittin-induced alterations in morphology and deformability of human red blood cells using quantitative phase imaging techniques

2016 ◽  
Author(s):  
Joonseok Hur ◽  
Kyoohyun Kim ◽  
SangYun Lee ◽  
HyunJoo Park ◽  
YongKeun Park
Keyword(s):  
Refractive Index ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Imaging Techniques ◽  
Phase Imaging ◽  
Quantitative Phase Imaging ◽  
Active Molecule ◽  
Quantitative Phase ◽  
Human Red Blood Cells ◽  
Phase Images

Here, the actions of melittin, the active molecule of apitoxin or bee venom, were investigated on human red blood cells (RBCs) using quantitative phase imaging techniques. High-resolution realtime 3-D refractive index (RI) measurements and dynamic 2-D phase images of individual melittin-bound RBCs enabled in-depth examination of melittin-induced biophysical alterations of the cells. From the measurements, morphological, biochemical, and mechanical alterations of the RBCs were analyzed quantitatively. Furthermore, leakage of haemoglobin (Hb) inside the RBCs at high melittin concentration was also investigated.

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