scholarly journals Three-dimensional Super-resolved Imaging of Paraffin-embedded Kidney Samples

2021 ◽  
Author(s):  
David Unnersjoe-Jess ◽  
Amer Ramdedovic ◽  
Martin Hoehne ◽  
Linus Butt ◽  
Felix C Koehler ◽  
...  
Keyword(s):  
New Technologies ◽  
Three Dimensional ◽  
Kidney Tissue ◽  
Confocal Imaging ◽  
Disease Assessment ◽  
Gold Standard Method ◽  
Filtration Barrier ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Monitoring Treatment

Diseases of the glomeruli, the renal filtration units, are a leading cause of progressive kidney disease. Assessment of the ultrastructure of podocytes at the glomerular filtration barrier is essential for diagnosing diverse disease entities, providing insight into the disease pathogenesis as well as monitoring treatment responses. New technologies, including super-resolved nanoscopy and expansion microscopy, as well as new sample preparation techniques, are starting to revolutionize imaging of biopsy specimens. However, our previous approaches for simple and fast three-dimensional imaging of optically cleared samples are to date not compatible with formalin fixed paraffin-embedded (FFPE) tissue, impeding application in clinical routine. Here we provide protocols that circumvent these limitations and allow for three dimensional STED and confocal imaging of FFPE kidney tissue with similar staining and image quality as compared to our previous approaches. This would increase the feasibility to implement these protocols in clinical routines, as FFPE is the gold standard method for storage of patient samples.

scholarly journals Three-Dimensional Super-Resolved Imaging of Paraffin-Embedded Kidney Samples

Kidney360 ◽  
2021 ◽  
pp. 10.34067/KID.0005882021
Author(s):  
David Unnersjö-Jess ◽  
Amer Ramdedovic ◽  
Martin Höhne ◽  
Linus Butt ◽  
Felix C. Koehler ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Kidney Tissue ◽  
Confocal Imaging ◽  
Disease Assessment ◽  
Preparation Methods ◽  
Gold Standard Method ◽  
Filtration Barrier ◽  
Disease Entities ◽  
Kidney Imaging ◽  
Monitoring Treatment

Background: Diseases of the glomeruli, the renal filtration units, are a leading cause of progressive kidney disease. Assessment of the ultrastructure of podocytes at the glomerular filtration barrier is essential for diagnosing diverse disease entities, providing insight into the disease pathogenesis as well as monitoring treatment responses. Methods: We here apply previously published sample preparation methods together with STED and confocal microscopy for resolving nanoscale podocyte substructure. The protocols are modified and optimized in order to be applied to samples which have been formalin fixed and paraffin-embedded (FFPE). Results: We successfully modify our protocols to allow for deep three-dimensional STED and confocal imaging of FFPE kidney tissue with similar staining and image quality as compared to our previous approaches. We further show that quantitative analysis can be applied to extract morphometrics of healthy and diseased samples from both mice and humans. Conclusions: The results from this study could increase the feasibility to implement optical kidney imaging protocols in clinical routines, as FFPE is the gold standard method for storage of patient samples.

scholarly journals Immunofluorescence-guided segmentation of three-dimensional features in micro-computed tomography datasets of human lung tissue

2021 ◽  
Vol 8 (11) ◽  
Author(s):  
Matthew J. Lawson ◽  
Orestis L. Katsamenis ◽  
David Chatelet ◽  
Aiman Alzetani ◽  
Oliver Larkin ◽  
...  
Keyword(s):  
Computed Tomography ◽  
Lung Tissue ◽  
Human Lung ◽  
Three Dimensional ◽  
Micro Computed Tomography ◽  
3D Segmentation ◽  
Human Lung Tissue ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed

Micro-computed tomography (µCT) provides non-destructive three-dimensional (3D) imaging of soft tissue microstructures. Specific features in µCT images can be identified using correlated two-dimensional (2D) histology images allowing manual segmentation. However, this is very time-consuming and requires specialist knowledge of the tissue and imaging modalities involved. Using a custom-designed µCT system optimized for imaging unstained formalin-fixed paraffin-embedded soft tissues, we imaged human lung tissue at isotropic voxel sizes less than 10 µm. Tissue sections were stained with haematoxylin and eosin or cytokeratin 18 in columnar airway epithelial cells using immunofluorescence (IF), as an exemplar of this workflow. Novel utilization of tissue autofluorescence allowed automatic alignment of 2D microscopy images to the 3D µCT data using scripted co-registration and automated image warping algorithms. Warped IF images, which were accurately aligned with the µCT datasets, allowed 3D segmentation of immunoreactive tissue microstructures in the human lung. Blood vessels were segmented semi-automatically using the co-registered µCT datasets. Correlating 2D IF and 3D µCT data enables accurate identification, localization and segmentation of features in fixed soft lung tissue. Our novel correlative imaging workflow provides faster and more automated 3D segmentation of µCT datasets. This is applicable to the huge range of formalin-fixed paraffin-embedded tissues held in biobanks and archives.

Optimized protocol for metabolomic and lipidomic profiling in formalin-fixed paraffin-embedded kidney tissue by LC-MS

2020 ◽  
Vol 1134 ◽  
pp. 125-135 ◽  
Author(s):  
Sylvia K. Neef ◽  
Stefan Winter ◽  
Ute Hofmann ◽  
Thomas E. Mürdter ◽  
Elke Schaeffeler ◽  
...  
Keyword(s):  
Kidney Tissue ◽  
Formalin Fixed Paraffin ◽  
Optimized Protocol ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed

Sequencing Analysis and Phylogenetic Tree of HPV Isolated from Breast Cancer Patients at Thi-Qar Province/Iraq

2020 ◽  
pp. 37-40
Keyword(s):  
Nucleotide Sequence ◽  
Phylogenetic Tree ◽  
Evolutionary Relationship ◽  
Nucleotide Sequencing ◽  
Sequencing Analysis ◽  
Breast Cancer Patients ◽  
The North ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
History Of

Genetic variety examination has demonstrated fundamental to the understanding of the epidemiological and developmental history of Papillomavirus (HPV), for the development of accurate diagnostic tests and for efficient vaccine design. The HPV nucleotide diversity has been investigated widely among high-risk HPV types. To make the nucleotide sequence of HPV and do the virus database in Thi-Qar province, and compare sequences of our isolates with previously described isolates from around the world and then draw its phylogenetic tree, this study done. A total of 6 breast formalin-fixed paraffin-embedded (FFPE) of the female patients were included in the study, divided as 4 FFPE malignant tumor and 2 FFPE of benign tumor. The PCR technique was implemented to detect the presence of HPV in breast tissue, and the real-time PCR used to determinant HPV genotypes, then determined a complete nucleotide sequence of HPV of L1 capsid gene, and draw its phylogenetic tree. The nucleotide sequencing finding detects a number of substitution mutation (SNPs) in (L1) gene, which have not been designated before, were identified once in this study population, and revealed that the HPV16 strains have the evolutionary relationship with the South African race, while, the HPV33 and HPV6 showing the evolutionary association with the North American and East Asian race, respectively.

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