scholarly journals Image-Based Method to Quantify Decellularization of Tissue Sections

2021 ◽  
Vol 22 (16) ◽  
pp. 8399
Author(s):  
Maria Narciso ◽  
Jorge Otero ◽  
Daniel Navajas ◽  
Ramon Farré ◽  
Isaac Almendros ◽  
...  
Keyword(s):  
Dna Content ◽  
Phase Contrast ◽  
Tissue Characterization ◽  
Dna Quantification ◽  
Local Entropy ◽  
Image Processing Algorithm ◽  
Tissue Sections ◽  
Contrast Image ◽  
Average Fluorescence ◽  
Tissue Digestion

Tissue decellularization is typically assessed through absorbance-based DNA quantification after tissue digestion. This method has several disadvantages, namely its destructive nature and inadequacy in experimental situations where tissue is scarce. Here, we present an image processing algorithm for quantitative analysis of DNA content in (de)cellularized tissues as a faster, simpler and more comprehensive alternative. Our method uses local entropy measurements of a phase contrast image to create a mask, which is then applied to corresponding nuclei labelled (UV) images to extract average fluorescence intensities as an estimate of DNA content. The method can be used on native or decellularized tissue to quantify DNA content, thus allowing quantitative assessment of decellularization procedures. We confirm that our new method yields results in line with those obtained using the standard DNA quantification method and that it is successful for both lung and heart tissues. We are also able to accurately obtain a timeline of decreasing DNA content with increased incubation time with a decellularizing agent. Finally, the identified masks can also be applied to additional fluorescence images of immunostained proteins such as collagen or elastin, thus allowing further image-based tissue characterization.

Characteristics of an Electrostatic Phase Plate

1972 ◽  
Vol 30 ◽  
pp. 618-619
Author(s):  
William Krakow ◽  
Benjamin Siegel
Keyword(s):  
Phase Contrast ◽  
Objective Lens ◽  
Phase Plate ◽  
Net Charge ◽  
Phase Plates ◽  
Beam Currents ◽  
Contrast Image ◽  
Bright Phase ◽  
Additional Phase Shift ◽  
Additional Phase

Unwin has used a metallized non-conducting thread in the back focal plane of the objective lens that stops out a portion of the unscattered beam, takes on a localized positive charge and thus produces an additional phase shift to give a different transfer function of the lens. Under the particular conditions Unwin used, the phase contrast image was shifted to bright phase contrast for optimum focus.We have investigated the characteristics of this type of electrostatic phase plate, both analytically and experimentally, as functions of the magnitude of charge and defocus. Phase plates have been constructed by using Wollaston wire to mount 0.25μ diameter platinum wires across apertures ranging from 50 to 200μ diameter and vapor depositing SiO and gold on the mounted wires to give them the desired charging characteristics. The net charge was varied by adjusting only the bias on the Wehnelt shield of the gun, and hence the beam currents and effective size of the source.

Evaluation method for imaging plate resolution by means of phase contrast transfer function

1995 ◽  
Vol 53 ◽  
pp. 662-663 ◽  
Author(s):  
T. Oikawa ◽  
H. Kosugi ◽  
F. Hosokawa ◽  
D. Shindo ◽  
M. Kersker
Keyword(s):  
Transfer Function ◽  
Phase Contrast ◽  
Evaluation Method ◽  
Amorphous Film ◽  
Imaging Plate ◽  
X Rays ◽  
Special Shape ◽  
Contrast Transfer Function ◽  
Contrast Image ◽  
Specimen Shape

Evaluation of the resolution of the Imaging Plate (IP) has been attempted by some methods. An evaluation method for IP resolution, which is not influenced by hard X-rays at higher accelerating voltages, was proposed previously by the present authors. This method, however, requires truoblesome experimental preperations partly because specially synthesized hematite was used as a specimen, and partly because a special shape of the specimen was used as a standard image. In this paper, a convenient evaluation method which is not infuenced by the specimen shape and image direction, is newly proposed. In this method, phase contrast images of thin amorphous film are used.Several diffraction rings are obtained by the Fourier transformation of a phase contrast image of thin amorphous film, taken at a large under focus. The rings show the spatial-frequency spectrum corresponding to the phase contrast transfer function (PCTF). The envelope function is obtained by connecting the peak intensities of the rings. The evelope function is offten used for evaluation of the instrument, because the function shows the performance of the electron microscope (EM).

Bonafide Substructure in the Core of Ferritin

1985 ◽  
Vol 43 ◽  
pp. 322-323
Author(s):  
William H. Massover
Keyword(s):  
Phase Contrast ◽  
Central Cavity ◽  
Amplitude Image ◽  
Ultrastructural Studies ◽  
The Core ◽  
Hydrated Ferric Oxide ◽  
Imaging Artifact ◽  
Contrast Image ◽  
Protein Shell ◽  
Regular Patterns

Each molecule of ferritin (d = 130Å) contains a core of iron surrounded by a 24-subunit protein shell. The amount of iron stored is variable and is present within the central cavity (d = 80Å) as a hydrated ferric oxide equivalent to the mineral, ferrihydrite. Many early ultrastructural studies of ferritin detected regular patterns of a multiparticulate substructure in the iron-rich core [e.g., 3,4], Each small particle was termed a “micelle“; a theory became widely accepted that a core consisted of up to six micelles positioned at the vertices of an octahedron. Other workers recognized that the apparent micelles were smaller or even disappeared if images were recorded closer to exact focus [e.g., 5]. In 1969, Haydon clearly established that the observed substructure was really an imaging artifact; each apparent micelle was only a dot in the underfocused phase contrast image of the supporting film superimposed on the amplitude image of the strongly scattering metal.

scholarly journals Implementation of Accurate and Fast DNA Cytometry by Confocal Microscopy in 3D

2005 ◽  
Vol 27 (4) ◽  
pp. 225-230
Author(s):  
Lennert S. Ploeger ◽  
André Huisman ◽  
Jurryt van der Gugten ◽  
Dionne M. van der Giezen ◽  
Jeroen A. M. Beliën ◽  
...  
Keyword(s):  
Genomic Instability ◽  
Confocal Laser Scanning Microscopy ◽  
Dna Content ◽  
Laser Scanning ◽  
Clinical Applications ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Confocal Laser ◽  
Dna Cytometry ◽  
Tissue Sections

Background: DNA cytometry is a powerful method for measuring genomic instability. Standard approaches that measure DNA content of isolated cells may induce selection bias and do not allow interpretation of genomic instability in the context of the tissue. Confocal Laser Scanning Microscopy (CLSM) provides the opportunity to perform 3D DNA content measurements on intact cells in thick histological sections. Because the technique is technically challenging and time consuming, only a small number of usually manually selected nuclei were analyzed in different studies, not allowing wide clinical evaluation. The aim of this study was to describe the conditions for accurate and fast 3D CLSM cytometry with a minimum of user interaction to arrive at sufficient throughput for pilot clinical applications. Methods: Nuclear DNA was stained in 14 μm thick tissue sections of normal liver and adrenal stained with either YOYO-1 iodide or TO-PRO-3 iodide. Different pre-treatment strategies were evaluated: boiling in citrate buffer (pH 6.0) followed by RNase application for 1 or 18 hours, or hydrolysis. The image stacks obtained with CLSM at microscope magnifications of ×40 or ×100 were analyzed off-line using in-house developed software for semi-automated 3D fluorescence quantitation. To avoid sectioned nuclei, the top and bottom of the stacks were identified from ZX and YZ projections. As a measure of histogram quality, the coefficient of variation (CV) of the diploid peak was assessed. Results: The lowest CV (10.3%) was achieved with a protocol without boiling, with 1 hour RNase treatment and TO-PRO-3 iodide staining, and a final image recording at ×60 or ×100 magnifications. A sample size of 300 nuclei was generally achievable. By filtering the set of automatically segmented nuclei based on volume, size and shape, followed by interactive removal of the few remaining faulty objects, a single measurement was completely analyzed in approximately 3 hours. Conclusions: The described methodology allows to obtain a largely unbiased sample of nuclei in thick tissue sections using 3D DNA cytometry by confocal laser scanning microscopy within an acceptable time frame for pilot clinical applications, and with a CV small enough to resolve smaller near diploid stemlines. This provides a suitable method for 3D DNA ploidy assessment of selected rare cells based on morphologic characteristics and of clinical samples that are too small to prepare adequate cell suspensions.

Experimental verification of phase contrast image synthesis system

1996 ◽  
Author(s):  
Jesper Glueckstad ◽  
Haruyoshi Toyoda ◽  
Narihiro Yoshida ◽  
Tamiki Takemori ◽  
Tsutomu Hara
Keyword(s):  
Experimental Verification ◽  
Phase Contrast ◽  
Image Synthesis ◽  
Phase Contrast Image ◽  
Synthesis System ◽  
Contrast Image

scholarly journals Automated imaging cytometry reveals dysplastic indices of colonic serrated adenomas

2020 ◽  
pp. FSO459
Author(s):  
Nicholas S Samel ◽  
Qin Huang ◽  
Hiroshi Mashimo
Keyword(s):  
Dna Content ◽  
Chromosome Instability ◽  
Neoplastic Progression ◽  
Hyperplastic Polyps ◽  
Dna Index ◽  
Tissue Sections ◽  
Automated Imaging ◽  
Imaging Cytometry ◽  
The Mean ◽  
Serrated Adenomas

Aim: Left-sided colonic serrated adenomas (L-SAs) were evaluated for aneuploidy using automated imaging cytometry to quantify DNA content and compared with normal colonic tissues (NCT), tubular adenomas (TA), left-sided hyperplastic polyps (L-HP) and adenocarcinomas. Materials & methods: We used standard paraffin-embedded Feulgen-stained tissue sections. Results: The mean DNA index (DI) of NCT was 0.95, L-HP was 1.08, TA was 1.22, L-SA was 1.11 and adenocarcinomas was 1.46. DI of L-SA was statistically higher than that of NCT, but not statistically different from L-HP. Conclusion: This study demonstrates that DIs correlate with the described neoplastic progression of L-SA, TA and L-SA compared with NCT and suggests that L-SA may be involved in a chromosome instability pathway of neoplastic progression.

scholarly journals Green Fluorescent Protein and Phase-Contrast Image Fusion via Generative Adversarial Networks

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Wei Tang ◽  
Yu Liu ◽  
Chao Zhang ◽  
Juan Cheng ◽  
Hu Peng ◽  
...  
Keyword(s):  
Phase Contrast ◽  
Fluorescent Protein ◽  
Structural Information ◽  
Generative Adversarial Networks ◽  
Phase Contrast Image ◽  
Functional Information ◽  
Adversarial Networks ◽  
Fused Image ◽  
Green Fluorescent ◽  
Contrast Image

In the field of cell and molecular biology, green fluorescent protein (GFP) images provide functional information embodying the molecular distribution of biological cells while phase-contrast images maintain structural information with high resolution. Fusion of GFP and phase-contrast images is of high significance to the study of subcellular localization, protein functional analysis, and genetic expression. This paper proposes a novel algorithm to fuse these two types of biological images via generative adversarial networks (GANs) by carefully taking their own characteristics into account. The fusion problem is modelled as an adversarial game between a generator and a discriminator. The generator aims to create a fused image that well extracts the functional information from the GFP image and the structural information from the phase-contrast image at the same time. The target of the discriminator is to further improve the overall similarity between the fused image and the phase-contrast image. Experimental results demonstrate that the proposed method can outperform several representative and state-of-the-art image fusion methods in terms of both visual quality and objective evaluation.

Sign in / Sign up

Export Citation Format

Share Document