scholarly journals Ex Vivo Confocal Fluorescence Microscopy for Rapid Evaluation of Tissues in Surgical Pathology Practice

2017 ◽  
Vol 142 (3) ◽  
pp. 396-401 ◽  
Author(s):  
Savitri Krishnamurthy ◽  
Andrea Cortes ◽  
Mirtha Lopez ◽  
Michael Wallace ◽  
Sharjeel Sabir ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Acridine Orange ◽  
Ex Vivo ◽  
Imaging Techniques ◽  
Surgical Pathology ◽  
Confocal Fluorescence Microscopy ◽  
Tissue Preparation ◽  
Tissue Sections ◽  
Confocal Fluorescence ◽  
Hematoxylin Eosin

Context.— Optical imaging techniques are currently available for imaging tissues without the need for any type of extensive tissue preparation. There are several applications for their potential use in surgical pathology practice. Objective.— To evaluate the feasibility of using a confocal fluorescence microscopy (CFM) platform for ex vivo examination of tissues obtained from surgical resections of breast, lung, kidney, and liver. Design.— Tissue fragments (0.5–1.0 cm) were immersed in 0.6 mM acridine orange for 6 seconds and imaged using a CFM platform at a 488-nm wavelength. The imaged tissues were subsequently fixed in formalin and processed routinely to generate hematoxylin-eosin–stained tissue sections. Mosaics of the grayscale CFM images were studied at different magnifications for recognition of the tissue and were compared with conventional histopathologic examination of hematoxylin-eosin tissue sections. Results.— We imaged 55 tissue fragments obtained from 16 breast (29%), 18 lung (33%), 14 kidney (25%), and 7 liver (13%) surgical excision specimens. Acridine orange labeled the nuclei, creating the contrast between nucleus and cytoplasm and thereby recapitulating the tissue architecture. We could obtain CFM images of good quality within 5 to 10 minutes that allowed recognition of the cytomorphologic details for categorization of the imaged tissue and were similar to histologic examination of hematoxylin-eosin tissue sections. Conclusions.— The ease and speed of acquisition of CFM images together with the resolution and resemblance of the CFM images to hematoxylin-eosin sections suggest that the CFM platform has excellent potential for use in surgical pathology practice.

scholarly journals Confocal Fluorescence Microscopy Platform Suitable for Rapid Evaluation of Small Fragments of Tissue in Surgical Pathology Practice

2018 ◽  
Vol 143 (3) ◽  
pp. 305-313 ◽  
Author(s):  
Savitri Krishnamurthy ◽  
Kechen Ban ◽  
Kenna Shaw ◽  
Gordon Mills ◽  
Rahul Sheth ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Predictive Value ◽  
Surgical Excision ◽  
Surgical Pathology ◽  
Confocal Fluorescence Microscopy ◽  
Rapid Evaluation ◽  
Tissue Sections ◽  
Confocal Fluorescence ◽  
Hematoxylin Eosin

Context.— Rapid advances in the fields of biophotonics, computer science, and instrumentation have allowed for high-resolution imaging of biologic tissues. Objective.— To evaluate the quality of images from an optimized confocal fluorescence microscopy (CFM) platform for rapid evaluation of small fragments of tissue, compared with hematoxylin-eosin staining. Design.— Tissue fragments (up to 1.0 × 0.3 cm) were stained with 0.6 mM acridine orange for 60 seconds and imaged using a CFM platform at 488-nm and 785-nm wavelength. The imaged tissues were then fixed in formalin and processed to generate hematoxylin-eosin–stained tissue sections. The quality of CFM images was scored on a scale of 0 to 3 on the basis of the percentage of the CFM images with recognizable tissue architecture (0, 0%; 1, <20%; 2, 20%–50%; 3, >50%). The diagnoses made using CFM images were compared with those made using histopathologic analysis of the hematoxylin-eosin–stained tissue sections. Results.— We imaged 118 tissue fragments obtained from 40 breast, 23 lung, 39 kidney, and 16 liver surgical excision specimens. We acquired CFM images in 2 to 3 minutes; 95.8% (113 of 118) of images showed a quality score of 3, and 4.2% (5 of 118) had a score of 2. We achieved a sensitivity of 95.5%, specificity of 97.3%, positive predictive value of 95.5%, and negative predictive value of 97.3%. Conclusions.— Our results demonstrate the suitability of the CFM platform for rapid and accurate evaluation of small tissue fragments in surgical pathology practice.

Smart imaging of acute lung injury: exploration of myeloperoxidase activity using in vivo endoscopic confocal fluorescence microscopy

2015 ◽  
Vol 309 (6) ◽  
pp. L543-L551 ◽  
Author(s):  
Frédéric Chagnon ◽  
Alexandra Bourgouin ◽  
Réjean Lebel ◽  
Marc-André Bonin ◽  
Eric Marsault ◽  
...  
Keyword(s):  
Acute Lung Injury ◽  
Lung Injury ◽  
Fluorescence Microscopy ◽  
Real Time ◽  
Ex Vivo ◽  
Polymorphonuclear Neutrophils ◽  
Confocal Fluorescence Microscopy ◽  
Myeloperoxidase Activity ◽  
Confocal Fluorescence

The pathophysiology of acute lung injury (ALI) is well characterized, but its real-time assessment at bedside remains a challenge. When patients do not improve after 1 wk despite supportive therapies, physicians have to consider open lung biopsy (OLB) to identify the process(es) at play. Sustained inflammation and inadequate repair are often observed in this context. OLB is neither easy to perform in a critical setting nor exempt from complications. Herein, we explore intravital endoscopic confocal fluorescence microscopy (ECFM) of the lung in vivo combined with the use of fluorescent smart probe(s) activated by myeloperoxidase (MPO). MPO is a granular enzyme expressed by polymorphonuclear neutrophils (PMNs) and alveolar macrophages (AMs), catalyzing the synthesis of hypoclorous acid, a by-product of hydrogen peroxide. Activation of these probes was first validated in vitro in relevant cells (i.e., AMs and PMNs) and on MPO-non-expressing cells (as negative controls) and then tested in vivo using three rat models of ALI and real-time intravital imaging with ECFM. Semiquantitative image analyses revealed that in vivo probe-related cellular/background fluorescence was associated with corresponding enhanced lung enzymatic activity and was partly prevented by specific MPO inhibition. Additional ex vivo phenotyping was performed, confirming that fluorescent cells were neutrophil elastase+(PMNs) or CD68+(AMs). This work is a first step toward “virtual biopsy” of ALI without OLB.

scholarly journals Fibered Confocal Fluorescence Microscopy for the Noninvasive Imaging of Langerhans Cells in Macaques

2017 ◽  
Vol 2017 ◽  
pp. 1-8 ◽  
Author(s):  
Biliana Todorova ◽  
Nina Salabert ◽  
Sabine Tricot ◽  
Raphaël Boisgard ◽  
Mélanie Rathaux ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Langerhans Cells ◽  
Ex Vivo ◽  
Confocal Fluorescence Microscopy ◽  
Noninvasive Technique ◽  
Hla Dr ◽  
Confocal Fluorescence ◽  
In Vivo Fluorescence Imaging ◽  
Fibered Confocal Fluorescence Microscopy

Purpose. We developed a new approach to visualize skin Langerhans cells by in vivo fluorescence imaging in nonhuman primates. Procedures. Macaques were intradermally injected with a monoclonal, fluorescently labeled antibody against HLA-DR molecule and were imaged for up to 5 days by fibered confocal microscopy (FCFM). Results. The network of skin Langerhans cells was visualized by in vivo fibered confocal fluorescence microscopy. Quantification of Langerhans cells revealed no changes to cell density with time. Ex vivo experiments confirmed that injected fluorescent HLA-DR antibody specifically targeted Langerhans cells in the epidermis. Conclusions. This study demonstrates the feasibility of single-cell, in vivo imaging as a noninvasive technique to track Langerhans cells in nontransgenic animals.

Utility of ex vivo confocal fluorescence microscopy for renal mass biopsy optimization

2021 ◽  
Vol 79 ◽  
pp. S817
Author(s):  
A.S. Valiquette ◽  
M. Marchioni ◽  
J. Rubio-Briones ◽  
A. Wong ◽  
A. Gomez-Ferrer ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Renal Mass ◽  
Ex Vivo ◽  
Confocal Fluorescence Microscopy ◽  
Confocal Fluorescence ◽  
Renal Mass Biopsy

Ex-vivo confocal fluorescence microscopy for rapid evaluation of renal core biopsy

2020 ◽  
Vol 72 (1) ◽  
Author(s):  
M. Carmen Mir ◽  
Bernardo Bancalari ◽  
Ana Calatrava ◽  
Juan Casanova ◽  
Jose-Luis Dominguez Escrig ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Core Biopsy ◽  
Ex Vivo ◽  
Confocal Fluorescence Microscopy ◽  
Rapid Evaluation ◽  
Confocal Fluorescence

scholarly journals Analysis of Polyhydroxyalkanoates Granules in Haloferax mediterranei by Double-Fluorescence Staining with Nile Red and SYBR Green by Confocal Fluorescence Microscopy

Polymers ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 1582
Author(s):  
Verónica Cánovas ◽  
Salvador Garcia-Chumillas ◽  
Fuensanta Monzó ◽  
Lorena Simó-Cabrera ◽  
Carmen Fernández-Ayuso ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Culture Media ◽  
Nile Red ◽  
Sybr Green ◽  
Confocal Fluorescence Microscopy ◽  
Fluorescence Staining ◽  
Haloferax Mediterranei ◽  
Optimized Protocol ◽  
Confocal Fluorescence ◽  
Pha Production

Haloferaxmediterranei is a haloarchaeon of high interest in biotechnology because it produces and mobilizes intracellular polyhydroxyalkanoate (PHA) granules during growth under stress conditions (limitation of phosphorous in the culture media), among other interesting metabolites (enzymes, carotenoids, etc.). The capability of PHA production by microbes can be monitored with the use of staining-based methods. However, the staining of haloarchaea cells is a challenging task; firstly, due to the high ionic strength of the medium, which is inappropriate for most of dyes, and secondly, due to the low permeability of the haloarchaea S-layer to macromolecules. In this work, Haloferax mediterranei is used as a halophilic archaeon model to describe an optimized protocol for the visualization and analysis of intracellular PHA granules in living cells. The method is based on double-fluorescence staining using Nile red and SYBR Green by confocal fluorescence microscopy. Thanks to this method, the capability of PHA production by new haloarchaea isolates could be easily monitored.

Sign in / Sign up

Export Citation Format

Share Document