The Effects of Illumination and Focal Distance on Light-Induced Fluorescence Images in vitro

2005 ◽  
Vol 40 (1) ◽  
pp. 73-76 ◽  
Author(s):  
I.A. Pretty ◽  
P.G. Ellwood ◽  
R.M. Davies ◽  
H.W. Worthington ◽  
R.P. Ellwood
Keyword(s):  
Focal Distance ◽  
Fluorescence Images

scholarly journals The Design and Preclinical Evaluation of a Single-Label Bimodal Nanobody Tracer for Image-Guided Surgery

Biomolecules ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 360
Author(s):  
Pieterjan Debie ◽  
Noemi B. Declerck ◽  
Danny van Willigen ◽  
Celine M. Huygen ◽  
Bieke De Sloovere ◽  
...  
Keyword(s):  
Single Molecule ◽  
Gamma Ray ◽  
Nuclear Imaging ◽  
Primary Amines ◽  
Resection Margins ◽  
Fluorescent Light ◽  
Single Structure ◽  
Fluorescence Images

Intraoperative guidance using targeted fluorescent tracers can potentially provide surgeons with real-time feedback on the presence of tumor tissue in resection margins. To overcome the limited depth penetration of fluorescent light, combining fluorescence with SPECT/CT imaging and/or gamma-ray tracing has been proposed. Here, we describe the design and preclinical validation of a novel bimodal nanobody-tracer, labeled using a “multifunctional single attachment point” (MSAP) label, integrating a Cy5 fluorophore and a diethylenetriaminepentaacetic acid (DTPA) chelator into a single structure. After conjugation of the bimodal MSAP to primary amines of the anti-HER2 nanobody 2Rs15d and 111In-labeling of DTPA, the tracer’s characteristics were evaluated in vitro. Subsequently, its biodistribution and tumor targeting were assessed by SPECT/CT and fluorescence imaging over 24 h. Finally, the tracer’s ability to identify small, disseminated tumor lesions was investigated in mice bearing HER2-overexpressing SKOV3.IP1 peritoneal lesions. [111In]In-MSAP.2Rs15d retained its affinity following conjugation and remained stable for 24 h. In vivo SPECT/CT and fluorescence images showed specific uptake in HER2-overexpressing tumors with low background. High tumor-to-muscle ratios were obtained at 1h p.i. and remained 19-fold on SPECT/CT and 3-fold on fluorescence images over 24 h. In the intraperitoneally disseminated model, the tracer allowed detection of larger lesions via nuclear imaging, while fluorescence enabled accurate removal of submillimeter lesions. Bimodal nuclear/fluorescent nanobody-tracers can thus be conveniently designed by conjugation of a single-molecule MSAP-reagent carrying a fluorophore and chelator for radioactive labeling. Such tracers hold promise for clinical applications.

scholarly journals How we discovered fluorescent speckle microscopy

2011 ◽  
Vol 22 (21) ◽  
pp. 3940-3942 ◽  
Author(s):  
E. D. Salmon ◽  
Clare M. Waterman
Keyword(s):  
Epithelial Cells ◽  
Fluorescence Intensity ◽  
Focal Adhesions ◽  
Time Lapse ◽  
Living Cells ◽  
Variable Intensity ◽  
Macromolecular Assemblies ◽  
Dynamic Assembly ◽  
Fluorescence Images

Fluorescent speckle microscopy (FSM) is a method for measuring the movements and dynamic assembly of macromolecular assemblies such as cytoskeletal filaments (e.g., microtubules and actin) or focal adhesions within large arrays in living cells or in preparations in vitro. The discovery of the method depended on recognizing the importance of unexpected fluorescence images of microtubules obtained by time-lapse recording of vertebrate epithelial cells in culture. In cells that were injected with fluorescent tubulin at ∼10% of the cytosol pool, microtubules typically appeared as smooth threads with a nearly constant fluorescence intensity. One day, when an unusually low concentration of fluorescent tubulin was injected into cells, the images from a sensitive cooled charge-coupled detector camera showed microtubules with an unusual “speckled” appearance—there were fluorescent dots with variable intensity and spacing along the microtubules. A first thought was that the speckles were an artifact. With further thought, we surmised that the speckles could be telling us something about stochastic association of tubulin dimers with the growing end of a microtubule. Numerous experiments confirmed the latter hypothesis. Subsequently the method we call FSM has proven to be very valuable. The speckles turned out not to be a meaningless artifact, but rather a serendipitous find.

scholarly journals AMES: Automated evaluation of sarcomere structures in cardiomyocytes

2021 ◽  
Author(s):  
Maximilian Hillemanns ◽  
Heiko Lemcke ◽  
Robert David ◽  
Thomas Martinetz ◽  
Markus Wolfien ◽  
...  
Keyword(s):  
Quality Control ◽  
Drug Testing ◽  
Cardiac Tissue ◽  
3D Models ◽  
Automated Evaluation ◽  
Cell Source ◽  
Unbiased Manner ◽  
Fluorescence Images ◽  
Induced Pluripotent

Background: Arrhythmias are severe cardiac diseases and lethal if untreated. To serve as an in vitro drug testing option for anti arrhythmic agents, cardiomyocytes are being generated in vitro from induced pluripotent stem cells (iPSCs). Unfortunately, these generated cardiomyocytes resemble fetal cardiac tissue rather than adult cardiomyocytes. An automated tool for an unbiased evaluation of cardiomyocytes would highly facilitate the establishment of new differentiation protocols to increase cellular maturity. Results: In this work, a novel deep learning-based approach for this task is presented and evaluated. Different convolutional neural networks (CNNs) including 2D and 3D models were trained on fluorescence images of human iPSC-derived cardiomyocytes, which were rated based on their sarcomere content (sarcomerisation) and the orientation of sarcomere filaments (directionality) beforehand by a domain expert. The CNNs were trained to perform classifications on sarcomerisation, directionality ratings, and cell source, including primary adult and differentiated cardiomyocytes. The best accuracies are reached by a 3D model with a classification accuracy of about 90 % for sarcomerisation classification, 63 % for directionality classification, and 80 % for cell source classification. The trained models were additionally evaluated using two explanatory algorithms, IGrad and Grad-CAM. The heatmaps computed by those explainability algorithms show that the important regions in the image occur inside the cell and at the cellular borders for the classifier, and, therefore, validate the calculated regions. Conclusion: In summary, we showed that cellular fluorescence images can be analyzed with CNNs and subsequently used to predict different states of sarcomere maturation. Our developed prediction tool AMES (https://github.com/maxhillemanns/AMES) can be used to make trustworthy predictions on the quality of a cardiomyocyte, which ultimately facilitates the optimized generation of cardiomyocytes from iPSCs and improves the quality control in an automated, unbiased manner. The applied workflow of testing different CNN models, adjusting parameters, and using a variety of explanatory algorithms can be easily transferred to further image based quality control, stratification, or analysis setups.

Seti Space Missions

1997 ◽  
Vol 161 ◽  
pp. 761-776 ◽  
Author(s):  
Claudio Maccone
Keyword(s):  
Electromagnetic Waves ◽  
Space Missions ◽  
Gravitational Lens ◽  
The Sun ◽  
Space Antenna ◽  
Focal Distance ◽  
Large Space ◽  
The Earth ◽  
Space Antennas ◽  
New Space

AbstractSETI from space is currently envisaged in three ways: i) by large space antennas orbiting the Earth that could be used for both VLBI and SETI (VSOP and RadioAstron missions), ii) by a radiotelescope inside the Saha far side Moon crater and an Earth-link antenna on the Mare Smythii near side plain. Such SETIMOON mission would require no astronaut work since a Tether, deployed in Moon orbit until the two antennas landed softly, would also be the cable connecting them. Alternatively, a data relay satellite orbiting the Earth-Moon Lagrangian pointL2would avoid the Earthlink antenna, iii) by a large space antenna put at the foci of the Sun gravitational lens: 1) for electromagnetic waves, the minimal focal distance is 550 Astronomical Units (AU) or 14 times beyond Pluto. One could use the huge radio magnifications of sources aligned to the Sun and spacecraft; 2) for gravitational waves and neutrinos, the focus lies between 22.45 and 29.59 AU (Uranus and Neptune orbits), with a flight time of less than 30 years. Two new space missions, of SETI interest if ET’s use neutrinos for communications, are proposed.

Ultrastructural Investigations of the Contractile Filaments of Amoebae

1974 ◽  
Vol 32 ◽  
pp. 188-189
Author(s):  
P.L. Moore
Keyword(s):  
Cytoplasmic Streaming ◽  
Three Dimensional ◽  
Freeze Fracture ◽  
Amoeboid Movement ◽  
Different Types ◽  
Chemical Conditions ◽  
Complete Interpretation

Previous freeze fracture results on the intact giant, amoeba Chaos carolinensis indicated the presence of a fibrillar arrangement of filaments within the cytoplasm. A complete interpretation of the three dimensional ultrastructure of these structures, and their possible role in amoeboid movement was not possible, since comparable results could not be obtained with conventional fixation of intact amoebae. Progress in interpreting the freeze fracture images of amoebae required a more thorough understanding of the different types of filaments present in amoebae, and of the ways in which they could be organized while remaining functional.The recent development of a calcium sensitive, demembranated, amoeboid model of Chaos carolinensis has made it possible to achieve a better understanding of such functional arrangements of amoeboid filaments. In these models the motility of demembranated cytoplasm can be controlled in vitro, and the chemical conditions necessary for contractility, and cytoplasmic streaming can be investigated. It is clear from these studies that “fibrils” exist in amoeboid models, and that they are capable of contracting along their length under conditions similar to those which cause contraction in vertebrate muscles.

In Vitro Induction of Crystals of MT Protein by Vinblastine-Colcemid in Mouse Spermatids

1974 ◽  
Vol 32 ◽  
pp. 132-133
Author(s):  
John J. Wolosewick ◽  
John H. D. Bryan
Keyword(s):  
Endoplasmic Reticulum ◽  
Smooth Endoplasmic Reticulum ◽  
Nuclear Ring ◽  
Rough Er ◽  
Distal Direction ◽  
In Vitro Induction

Early in spermiogenesis the manchette is rapidly assembled in a distal direction from the nuclear-ring-densities. The association of vesicles of smooth endoplasmic reticulum (SER) and the manchette microtubules (MTS) has been reported. In the mouse, osmophilic densities at the distal ends of the manchette are the organizing centers (MTOCS), and are associated with the SER. Rapid MT assembly and the lack of rough ER suggests that there is an existing pool of MT protein. Colcemid potentiates the reaction of vinblastine with tubulin and was used in this investigation to detect this protein.

Induction and Differentiation of Dental Epithelium in Vivo and in Vitro

1982 ◽  
Vol 40 ◽  
pp. 142-145
Author(s):  
E. J. Kollar
Keyword(s):  
Connective Tissue ◽  
Oral Mucosa ◽  
Basal Lamina ◽  
Functional Derivatives ◽  
Specific Source ◽  
Dental Epithelium ◽  
Connective Tissue Stroma

The differentiation and maintenance of many specialized epithelial structures are dependent on the underlying connective tissue stroma and on an intact basal lamina. These requirements are especially stringent in the development and maintenance of the skin and oral mucosa. The keratinization patterns of thin or thick cornified layers as well as the appearance of specialized functional derivatives such as hair and teeth can be correlated with the specific source of stroma which supports these differentiated expressions.

Immunoenzymatic demonstration of the simultaneous presence of transferrin, hemopexin and albumin in a same hepatocyte and their ultrastructural localization

1978 ◽  
Vol 36 (2) ◽  
pp. 88-89
Author(s):  
M. Kraemer ◽  
J. Foucrier ◽  
J. Vassy ◽  
M.T. Chalumeau
Keyword(s):  
Plasma Proteins ◽  
Rat Hepatocytes ◽  
Ultrastructural Localization ◽  
Carrier Proteins ◽  
Normal Cells ◽  
Adult Rat ◽  
Adult Rat Hepatocytes ◽  
Monospecific Antisera ◽  
Different Levels

Some authors using immunofluorescent techniques had already suggested that some hepatocytes are able to synthetize several plasma proteins. In vitro studies on normal cells or on cells issued of murine hepatomas raise the same conclusion. These works could be indications of an hepatocyte functionnal non-specialization, meanwhile the authors never give direct topographic proofs suitable with this hypothesis.The use of immunoenzymatic techniques after obtention of monospecific antisera had seemed to us useful to bring forward a better knowledge of this problem. We have studied three carrier proteins (transferrin = Tf, hemopexin = Hx, albumin = Alb) operating at different levels in iron metabolism by demonstrating and localizing the adult rat hepatocytes involved in their synthesis.Immunological, histological and ultrastructural methods have been described in a previous work.

Sem Studies of Co-Cr-Mo Surgical Implant Alloy Corrosion Behavior

1982 ◽  
Vol 40 ◽  
pp. 520-521
Author(s):  
Ann Chidester Van Orden ◽  
John L. Chidester ◽  
Anna C. Fraker ◽  
Pei Sung
Keyword(s):  
Electron Microscopy ◽  
Corrosion Behavior ◽  
Anodic Polarization ◽  
Saline Solution ◽  
Saturated Calomel Electrode ◽  
Physiological Saline ◽  
X Ray ◽  
Physiological Saline Solution ◽  
Scanning Electron

The influence of small variations in the composition on the corrosion behavior of Co-Cr-Mo alloys has been studied using scanning electron microscopy (SEM), energy dispersive x-ray analysis (EDX), and electrochemical measurements. SEM and EDX data were correlated with data from in vitro corrosion measurements involving repassivation and also potentiostatic anodic polarization measurements. Specimens studied included the four alloys shown in Table 1. Corrosion tests were conducted in Hanks' physiological saline solution which has a pH of 7.4 and was held at a temperature of 37°C. Specimens were mechanically polished to a surface finish with 0.05 µm A1203, then exposed to the solution and anodically polarized at a rate of 0.006 v/min. All voltages were measured vs. the saturated calomel electrode (s.c.e.).. Specimens had breakdown potentials near 0.47V vs. s.c.e.

Ultrastructural Correlations between Clonal Human Tumor Colonies and in Vivo Neoplasms

1982 ◽  
Vol 40 ◽  
pp. 210-211
Author(s):  
M.J. Murphy ◽  
R.R. Price ◽  
J.C. Sloman
Keyword(s):  
Cultured Cells ◽  
Human Tumor ◽  
Cell Type ◽  
Tumor Mass ◽  
Initial Cell ◽  
Cloning Assay ◽  
Human Tumor Cloning ◽  
The Relationship

The in vitro human tumor cloning assay originally described by Salmon and Hamburger has been applied recently to the investigation of differential anti-tumor drug sensitivities over a broad range of human neoplasms. A major problem in the acceptance of this technique has been the question of the relationship between the cultured cells and the original patient tumor, i.e., whether the colonies that develop derive from the neoplasm or from some other cell type within the initial cell population. A study of the ultrastructural morphology of the cultured cells vs. patient tumor has therefore been undertaken to resolve this question. Direct correlation was assured by division of a common tumor mass at surgical resection, one biopsy being fixed for TEM studies, the second being rapidly transported to the laboratory for culture.

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