3D visualization in multifocus fluorescence microscopy

2019 ◽  
Author(s):  
Julia R. Alonso ◽  
Alejandro Silva ◽  
Miguel Arocena
Keyword(s):  
Fluorescence Microscopy ◽  
3D Visualization

scholarly journals Light sheet fluorescence microscopy as a new method for unbiased three-dimensional analysis of vascular injury

2020 ◽  
Author(s):  
Nicholas E Buglak ◽  
Jennifer Lucitti ◽  
Pablo Ariel ◽  
Sophie Maiocchi ◽  
Francis J Miller ◽  
...  
Keyword(s):  
Carotid Artery ◽  
Fluorescence Microscopy ◽  
Vascular Injury ◽  
Histological Analysis ◽  
3D Visualization ◽  
Three Dimensional ◽  
Light Sheet ◽  
3D Analysis ◽  
Preclinical Models ◽  
Light Sheet Fluorescence Microscopy

Abstract Aims Assessment of preclinical models of vascular disease is paramount in the successful translation of novel treatments. The results of these models have traditionally relied on two-dimensional (2D) histological methodologies. Light sheet fluorescence microscopy (LSFM) is an imaging platform that allows for three-dimensional (3D) visualization of whole organs and tissues. In this study, we describe an improved methodological approach utilizing LSFM for imaging of preclinical vascular injury models while minimizing analysis bias. Methods and results The rat carotid artery segmental pressure-controlled balloon injury and mouse carotid artery ligation injury were performed. Arteries were harvested and processed for LSFM imaging and 3D analysis, as well as for 2D area histological analysis. Artery processing for LSFM imaging did not induce vessel shrinkage or expansion and was reversible by rehydrating the artery, allowing for subsequent sectioning and histological staining a posteriori. By generating a volumetric visualization along the length of the arteries, LSFM imaging provided different analysis modalities including volumetric, area, and radial parameters. Thus, LSFM-imaged arteries provided more precise measurements compared to classic histological analysis. Furthermore, LSFM provided additional information as compared to 2D analysis in demonstrating remodelling of the arterial media in regions of hyperplasia and periadventitial neovascularization around the ligated mouse artery. Conclusion LSFM provides a novel and robust 3D imaging platform for visualizing and quantifying arterial injury in preclinical models. When compared with classic histology, LSFM outperformed traditional methods in precision and quantitative capabilities. LSFM allows for more comprehensive quantitation as compared to traditional histological methodologies, while minimizing user bias associated with area analysis of alternating, 2D histological artery cross-sections.

Application of FRAP and digitized fluorescence microscopy to problems in cell membrane dynamics

1988 ◽  
Vol 46 ◽  
pp. 118-119
Author(s):  
K. Jacobson ◽  
A. Ishihara ◽  
B. Holifield ◽  
F. Zhang
Keyword(s):  
Fluorescence Microscopy ◽  
Cell Biology ◽  
Extended Period ◽  
Dynamic Structure ◽  
Living Cells ◽  
Membrane Dynamics ◽  
Molecular Cell Biology ◽  
Image Averaging ◽  
Single Living Cells ◽  
Single Living

Our laboratory is concerned with understanding the dynamic structure of the plasma membrane with particular reference to the movement of membrane constituents during cell locomotion. In addition to the standard tools of molecular cell biology, we employ both fluorescence recovery after photo- bleaching (FRAP) and digitized fluorescence microscopy (DFM) to investigate individual cells. FRAP allows the measurement of translational mobility of membrane and cytoplasmic molecules in small regions of single, living cells. DFM is really a new form of light microscopy in that the distribution of individual classes of ions, molecules, and macromolecules can be followed in single, living cells. By employing fluorescent antibodies to defined antigens or fluorescent analogs of cellular constituents as well as ultrasensitive, electronic image detectors and video image averaging to improve signal to noise, fluorescent images of living cells can be acquired over an extended period without significant fading and loss of cell viability.

Scanning x-ray fluorescence microscopy and principal component analysis

1992 ◽  
Vol 50 (2) ◽  
pp. 1752-1753
Author(s):  
Brian Cross
Keyword(s):  
Principal Component Analysis ◽  
Fluorescence Microscopy ◽  
Spatial Resolution ◽  
High Speed ◽  
Image Acquisition ◽  
Principal Component ◽  
Fluorescence Microscope ◽  
Acquisition Rate ◽  
X Ray ◽  
Speed Up

A relatively new entry, in the field of microscopy, is the Scanning X-Ray Fluorescence Microscope (SXRFM). Using this type of instrument (e.g. Kevex Omicron X-ray Microprobe), one can obtain multiple elemental x-ray images, from the analysis of materials which show heterogeneity. The SXRFM obtains images by collimating an x-ray beam (e.g. 100 μm diameter), and then scanning the sample with a high-speed x-y stage. To speed up the image acquisition, data is acquired "on-the-fly" by slew-scanning the stage along the x-axis, like a TV or SEM scan. To reduce the overhead from "fly-back," the images can be acquired by bi-directional scanning of the x-axis. This results in very little overhead with the re-positioning of the sample stage. The image acquisition rate is dominated by the x-ray acquisition rate. Therefore, the total x-ray image acquisition rate, using the SXRFM, is very comparable to an SEM. Although the x-ray spatial resolution of the SXRFM is worse than an SEM (say 100 vs. 2 μm), there are several other advantages.

Two-photon excitation fluorescence microscopy in living systems

1993 ◽  
Vol 51 ◽  
pp. 154-155 ◽  
Author(s):  
David W. Piston
Keyword(s):  
Confocal Microscopy ◽  
Fluorescence Microscopy ◽  
Singlet State ◽  
Excited Electronic State ◽  
Input Power ◽  
Two Photon ◽  
Simultaneous Absorption ◽  
Photon Excitation ◽  
Very High ◽  
Two Photon Excitation

Two-photon excitation fluorescence microscopy provides attractive advantages over confocal microscopy for three-dimensionally resolved fluorescence imaging. Two-photon excitation arises from the simultaneous absorption of two photons in a single quantitized event whose probability is proportional to the square of the instantaneous intensity. For example, two red photons can cause the transition to an excited electronic state normally reached by absorption in the ultraviolet. In our fluorescence experiments, the final excited state is the same singlet state that is populated during a conventional fluorescence experiment. Thus, the fluorophore exhibits the same emission properties (e.g. wavelength shifts, environmental sensitivity) used in typical biological microscopy studies. In practice, two-photon excitation is made possible by the very high local instantaneous intensity provided by a combination of diffraction-limited focusing of a single laser beam in the microscope and the temporal concentration of 100 femtosecond pulses generated by a mode-locked laser. Resultant peak excitation intensities are 106 times greater than the CW intensities used in confocal microscopy, but the pulse duty cycle of 10−5 maintains the average input power on the order of 10 mW, only slightly greater than the power normally used in confocal microscopy.

Localization of Immune Complexes in the Basement Membrane of the Ductuli Efferentes of the Rhesus Monkey

1980 ◽  
Vol 38 ◽  
pp. 456-457
Author(s):  
D. Marsh
Keyword(s):  
Basement Membrane ◽  
Fluorescence Microscopy ◽  
Rhesus Monkey ◽  
Immune Complex ◽  
Immune Complexes ◽  
Vas Deferens ◽  
Frozen Sections ◽  
Efferent Ducts ◽  
Complex Deposition ◽  
Sperm Antibodies

As a result of vasectomy, spermatozoa are confined to the epididymis and vas deferens, where they degenerate, releasing antigens that enter the circulation or are engulfed by macrophages. Multiple antigens of the sperm can elicit production of autoantibodies; circulating anti-sperm antibodies are found in a large percentage of vasectomized men, indicating the immunogenicity of the sperm. The increased prevalence of macrophages in the liomen of the rhesus monkey testicular efferent ducts after vasectomy led to further study of this region. Frozen sections were used for evaluation of immunopathological status by fluorescence microscopy with fluorescein-conjugated antibody. Subsequent granular deposits of immune complexes were revealed by positive immunofluorescence staining for complement. The immune complex deposition in the basement membrane surrounding the efferent ducts implies that this region is involved in antigen leakage (Fig. 1).

Alternative 2D and 3D Visualization Methods for Microsurgery: Posture, Performance, and Discomfort Analysis

2011 ◽  
Author(s):  
Denny Yu ◽  
Michael Sackllah ◽  
Charles Woolley ◽  
Steven Kasten ◽  
Thomas J. Armstrong
Keyword(s):  
3D Visualization ◽  
2D And 3D

INFLUENCES OF 3D VISUALIZATION AND FREE-POINT-VIEW TECHNOLOGYIES ON SPORTS VIEWING EXPERIENCES

2016 ◽  
Author(s):  
Jongho Kim ◽  
Yeayoung Noh ◽  
Yoonji Ryu ◽  
Carolyn Yoonhee Park ◽  
Choong Hoon Lim ◽  
...  
Keyword(s):  
3D Visualization
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