scholarly journals Extended Dual-Focus Microscopy for Ratiometric-Based 3D Movement Tracking

2020 ◽  
Vol 10 (18) ◽  
pp. 6243
Author(s):  
Seohyun Lee ◽  
Hyuno Kim ◽  
Hideo Higuchi
Keyword(s):  
Drug Delivery ◽  
High Resolution ◽  
Optical Axis ◽  
Three Dimensional ◽  
Virus Transmission ◽  
Living Cells ◽  
Depth Of Field ◽  
Objective Lens ◽  
Accurate Localization ◽  
Movement Tracking

Imaging the three-dimensional movement of small organelles in living cells can provide key information for the dynamics of drug delivery and virus transmission in biomedical disciplines. To stably monitor such intracellular motion using microscope, long depth of field along optical axis and accurate three-dimensional tracking are simultaneously required. In the present work, we suggest an extended dual-focus optics microscopy system by combining a bifocal plane imaging scheme and objective lens oscillation, which enables accurate localization for a long axial range. The proposed system exploits high-resolution functionality by concatenating partial calibration result acquired each axial imaging level, maintaining the practical advantages of ratiometric method.

400 Kv Electron Cryo-Microscopic Imaging Of Large Icosahedral Viruses Towards Atomic Resolution

1999 ◽  
Vol 5 (S2) ◽  
pp. 186-187
Author(s):  
Joanita Jakarta ◽  
Wah Chiu
Keyword(s):  
High Resolution ◽  
Three Dimensional ◽  
Clinical Manifestations ◽  
Image Data ◽  
Chromatic Aberration ◽  
Dimensional Structure ◽  
Depth Of Field ◽  
Periodic Arrays ◽  
Icosahedral Viruses ◽  
Simplex Virus

Three-dimensional structure studies provide important information about the organization of macromolecules, often revealing biological mechanisms and protein structure-function relationships. 400 KV electron cryo-microscopy is an emerging technology that is proving to be a powerful tool for studying the structures of large macromolecular assemblies that are often not tractable using other techniques. Its large depth of field makes it well-suited for imaging large objects to high resolution. In addition, a high accelerating voltage minimizes chromatic aberration yielding images of higher contrast. Recently a 400 KV electron cryo-microscope has been used to image periodic arrays of tubulin to 3.5 Å and single particles at somewhat lower resolutions (13 Å) providing practical demonstrations of its usefulness in modern structural biology. In this paper we present high resolution image data of two large icosahedral viruses: herpes simplex virus IB nucleocapsid (HSV IB) and rice dwarf virus (RDV). Human herpes virus (HSV) is associated with a spectrum of diseases ranging from cold sores to more severe clinical manifestations such as mental retardation.

scholarly journals Optical Projection Tomography Using a Commercial Microfluidic System

Micromachines ◽  
2020 ◽  
Vol 11 (3) ◽  
pp. 293
Author(s):  
Wenhao Du ◽  
Cheng Fei ◽  
Junliang Liu ◽  
Yongfu Li ◽  
Zhaojun Liu ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Numerical Aperture ◽  
Microfluidic System ◽  
Depth Of Field ◽  
Objective Lens ◽  
Flow Mode ◽  
Wide Field ◽  
Optical Projection Tomography ◽  
Microfluidic Systems ◽  
Optical Projection

Optical projection tomography (OPT) is the direct optical equivalent of X-ray computed tomography (CT). To obtain a larger depth of field, traditional OPT usually decreases the numerical aperture (NA) of the objective lens to decrease the resolution of the image. So, there is a trade-off between sample size and resolution. Commercial microfluidic systems can observe a sample in flow mode. In this paper, an OPT instrument is constructed to observe samples. The OPT instrument is combined with commercial microfluidic systems to obtain a three-dimensional and time (3D + T)/four-dimensional (4D) video of the sample. “Focal plane scanning” is also used to increase the images’ depth of field. A series of two-dimensional (2D) images in different focal planes was observed and compared with images simulated using our program. Our work dynamically monitors 3D OPT images. Commercial microfluidic systems simulate blood flow, which has potential application in blood monitoring and intelligent drug delivery platforms. We design an OPT adaptor to perform OPT on a commercial wide-field inverted microscope (Olympusix81). Images in different focal planes are observed and analyzed. Using a commercial microfluidic system, a video is also acquired to record motion pictures of samples at different flow rates. To our knowledge, this is the first time an OPT setup has been combined with a microfluidic system.

scholarly journals Connexin-specific distribution within gap junctions revealed in living cells

2000 ◽  
Vol 113 (22) ◽  
pp. 4109-4120 ◽  
Author(s):  
M.M. Falk
Keyword(s):  
High Resolution ◽  
Gap Junctions ◽  
Gap Junction ◽  
Fluorescent Protein ◽  
Three Dimensional ◽  
Time Lapse ◽  
Living Cells ◽  
Dye Transfer ◽  
Channel Distribution ◽  
Specific Distribution

To study the organization of gap junctions in living cells, the connexin isotypes alpha(1)(Cx43), beta(1)(Cx32) and beta(2)(Cx26) were tagged with the autofluorescent tracer green fluorescent protein (GFP) and its cyan (CFP) and yellow (YFP) color variants. The cellular fate of the tagged connexins was followed by high-resolution fluorescence deconvolution microscopy and time-lapse imaging. Comprehensive analyses demonstrated that the tagged channels were functional as monitored by dye transfer, even under conditions where the channels were assembled solely from tagged connexins. High-resolution images revealed a detailed structural organization, and volume reconstructions provided a three-dimensional view of entire gap junction plaques. Specifically, deconvolved dual-color images of gap junction plaques assembled from CFP- and YFP-tagged connexins revealed that different connexin isotypes gathered within the same plaques. Connexins either codistributed homogeneously throughout the plaque, or each connexin isotype segregated into well-separated domains. The studies demonstrate that the mode of channel distribution strictly depends on the connexin isotypes. Based on previous studies on the synthesis and assembly of connexins I suggest that channel distribution is regulated by intrinsic connexin isotype specific signals.

HREM autotuning on crystalline materials

1995 ◽  
Vol 53 ◽  
pp. 24-25
Author(s):  
M. Pan ◽  
O.L. Krivanek
Keyword(s):  
High Resolution ◽  
Amorphous Materials ◽  
Optical Axis ◽  
Amorphous Layer ◽  
Objective Lens ◽  
Crystalline Materials ◽  
Crystalline Sample ◽  
Resolution Electron ◽  
High Resolution Electron Microscope ◽  
Electron Microscopes

Complete autotuning of a high resolution electron microscope has been well established. It performs the following tasks: align the electron beam along the true electron-optical axis of objective lens (autoalignment), correct the astigmatism (autostigmation), and set the defocus to a user defined value (autofocus). It can also characterize the coefficient of 3-fold astigmatism while performing the autoalignment. Based on diffractogram analysis current HREM autotuning algorithm only works on amorphous materials. In reality, however, most of the HREM practice is performed on crystalline materials. Therefore it is highly desirable to extend the current HREM autotuning algorithm to crystalline specimens. In this abstract we report preliminary studies on attempting to analyze diffractograms from a mix of crystalline and amorphous materials.For crystalline specimens observed in most high resolution electron microscopes, except under UHV conditions, there is typically a thin layer of amorphous contamination due to either sample preparation or poor vacuum conditions. This amorphous layer can be easily seen at the edge of a crystalline sample in the microscope.

Light Microscopy Image Collection: Confocal, Widefield and Deconvolution

1997 ◽  
Vol 3 (S2) ◽  
pp. 371-372
Author(s):  
J. N. Turner ◽  
D. H. Szarowski ◽  
B. Roysam ◽  
T. J. Holmes
Keyword(s):  
Light Microscopy ◽  
Three Dimensional ◽  
Image Plane ◽  
Depth Of Field ◽  
Objective Lens ◽  
Biological Applications ◽  
Optical Sectioning ◽  
Microscopy Image ◽  
And Function ◽  
Image Collection

Light microscopy instrumentation and biological applications continue to expand rapidly, and an important aspect of this expansion is three-dimensional (3-D) imaging and image analysis. For many biological specimens, optical sectioning, i.e. collecting images at a sequence of depths under controlled conditions, provides true 3-D data through the entire specimen. This is especially important for specimens whose thickness exceeds the depth-of-field of the microscope objective lens. This sequence of optical sections is the basis for 3-D image reconstruction providing information from all three specimen dimensions instead of just the traditional two in the image plane of the microscope. The analysis of images in 3-D provides insights into the structure and function of biological specimens that are not available through other means. However, 3-D microscopy also presents additional choices for image collection. The first of which is whether to use widefield or confocal microscopy and the second is whether to utilize digital deblurring or deconvolution methods.

scholarly journals Two-Dimensional and Three-Dimensional Single Particle Tracking of Upconverting Nanoparticles in Living Cells

2019 ◽  
Vol 20 (6) ◽  
pp. 1424 ◽  
Author(s):  
Kyujin Shin ◽  
Yo Song ◽  
Yeongchang Goh ◽  
Kang Lee
Keyword(s):  
Particle Tracking ◽  
Single Particle ◽  
Near Infrared ◽  
Imaging Techniques ◽  
Three Dimensional ◽  
Living Cells ◽  
Single Particle Tracking ◽  
Infrared Light ◽  
Objective Lens ◽  
Two Dimensional

Lanthanide-doped upconversion nanoparticles (UCNPs) are inorganic nanomaterials in which the lanthanide cations embedded in the host matrix can convert incident near-infrared light to visible or ultraviolet light. These particles are often used for long-term and real-time imaging because they are extremely stable even when subjected to continuous irradiation for a long time. It is now possible to image their movement at the single particle level with a scale of a few nanometers and track their trajectories as a function of time with a scale of a few microseconds. Such UCNP-based single-particle tracking (SPT) technology provides information about the intracellular structures and dynamics in living cells. Thus far, most imaging techniques have been built on fluorescence microscopic techniques (epifluorescence, total internal reflection, etc.). However, two-dimensional (2D) images obtained using these techniques are limited in only being able to visualize those on the focal planes of the objective lens. On the contrary, if three-dimensional (3D) structures and dynamics are known, deeper insights into the biology of the thick cells and tissues can be obtained. In this review, we introduce the status of the fluorescence imaging techniques, discuss the mathematical description of SPT, and outline the past few studies using UCNPs as imaging probes or biologically functionalized carriers.

High Resolution Goniometer Stage for Electron Microscope

1971 ◽  
Vol 29 ◽  
pp. 42-43
Author(s):  
A. Mikaziri ◽  
S. Ohomori ◽  
T. Yamamoto
Keyword(s):  
Thermal Stability ◽  
Electron Microscope ◽  
High Resolution ◽  
Optical Axis ◽  
Resolving Power ◽  
Objective Lens ◽  
Thermal Drift ◽  
Power Stability ◽  
Supporting System

The performance of a goniometer stage to an electron microscope may be evaluated from several view points, such as operational easy, stability and resolving power. Stability is the fundamental, however. As is often experienced, the incorporation of a specimen tilting mechanism into the specimen chamber causes a functional unstability of the instrument and also causes inconvenience in the design of the objective lens. The top entry goniometer stage has an advantage in mechanical and thermal stability, since the specimen and specimen shifting stage are symmetrically supported around the optical axis. This supporting system is the main difference from that of the side entry goniometer stage. In the latter, the vibration and thermal drift of the specimen is unavoidable, in principle, due to an unbalanced specimen support.We have recently developed a top entry goniometer for the JEM-100B Electron Microscope, taking these points into account to meet the demand for high resolving power: this goniometer is of double tilt type, permitting each axis to tilt by ±30° and enabling the specimen to tilt around some composite tilting axis by ±40°.

scholarly journals High-Resolution Three-Dimensional Computed Tomography for Assessing Complications Related to Intrathecal Drug Delivery

2016 ◽  
Vol 5;19 (5;19) ◽  
pp. E775-E780
Author(s):  
Matthias Morgalla
Keyword(s):  
Computed Tomography ◽  
Drug Delivery ◽  
Chronic Pain ◽  
High Resolution ◽  
Volume Rendering ◽  
Predictive Value ◽  
Three Dimensional ◽  
3D Ct ◽  
Intrathecal Drug Delivery ◽  
Axial Ct

Background: The assessment of the functionality of intrathecal drug delivery (IDD) systems remains difficult and time-consuming. Catheter-related problems are still very common, and sometimes difficult to diagnose. Objectives: The aim of the present study is to investigate the accuracy of high-resolution three-dimensional computed tomography (CT) in order to detect catheter-related pump dysfunction. Study Design: An observational, retrospective investigation. Setting: Academic medical center in Germany. Methods: We used high-resolution three dimensional (3D) computed tomography with volume rendering technique (VRT) or fluoroscopy and conventional axial-CT to assess IDDrelated complications in 51 patients from our institution who had IDD systems implanted for the treatment of chronic pain or spasticity. Results: Twelve patients (23.5%) presented a total of 22 complications. The main type of complication in our series was catheter-related (50%), followed by pump failure, infection, and inappropriate refilling. Fluoroscopy and conventional CT were used in 12 cases. High-resolution 3D CT VRT scan was used in 35 instances with suspected yet unclear complications. Using 3D-CT (VRT) the sensitivity was 58.93% – 100% (CI 95%) and the specificity 87.54% – 100% (CI 95%).The positive predictive value was 58.93% – 100% (CI 95%) and the negative predictive value: 87.54% – 100% (CI 95%). Fluoroscopy and axial CT as a combined diagnostic tool had a sensitivity of 8.3% – 91.7% (CI 95%) and a specificity of 62.9% – 100% (CI 95%). The positive predictive value was 19.29% – 100% (CI 95%) and the negative predictive value: 44.43% – 96.89% (CI 95%). Limitations: This study is limited by its observational design and the small number of cases. Conclusion: High-resolution 3D CT VRT is a non- invasive method that can identify IDDrelated complications with more precision than axial CT and fluoroscopy. Key words: Volume rendering technique (VRT), intrathecal drug delivery systems, highresolution 3D computed tomography, complications, spasticity, chronic pain, cost effectiveness, fluoroscopy

Why Do We Need to Use Three-Dimensional (3D) Fourier Transform (FT) Analysis to Evaluate a High-Performance Transmission Electron Microscope (TEM)?

2016 ◽  
Vol 22 (5) ◽  
pp. 971-980 ◽  
Author(s):  
Kazuo Ishizuka ◽  
Koji Kimoto
Keyword(s):  
Fourier Transform ◽  
Electron Microscope ◽  
High Resolution ◽  
Spherical Aberration ◽  
Three Dimensional ◽  
Temporal Coherence ◽  
Necessary Condition ◽  
Objective Lens ◽  
Transmission Electron ◽  
Ewald Sphere

AbstractThe resolution of high-resolution transmission electron microscopes (TEM) has been improved down to subangstrom levels by correcting the spherical aberration (Cs) of the objective lens, and the information limit is thus determined mainly by partial temporal coherence. As a traditional Young’s fringe test does not reveal the true information limit for an ultra-high-resolution electron microscope, new methods to evaluate temporal coherence have been proposed based on a tilted-beam diffractogram. However, the diffractogram analysis cannot be applied when the nonlinear contribution becomes significant. Therefore, we have proposed a method based on the three-dimensional (3D) Fourier transform (FT) of through-focus TEM images, and evaluated the performance of someCs-corrected TEMs at lower voltages. In this report, we generalize the 3D FT analysis and derive the 3D transmission cross-coefficient. The profound difference of the 3D FT analysis from the diffractogram analysis is its capability to extract linear image information from the image intensity, and further to evaluate two linear image contributions separately on the Ewald sphere envelopes. Therefore, contrary to the diffractogram analysis the 3D FT analysis can work with a strong scattering object. This is the necessary condition if we want to directly observe the linear image transfer down to a few tens of picometer.

Isotropic high-resolution three-dimensional confocal micro-rotation imaging for non-adherent living cells

2009 ◽  
Vol 233 (3) ◽  
pp. 404-416 ◽  
Author(s):  
B. LE SAUX ◽  
B. CHALMOND ◽  
Y. YU ◽  
A. TROUVÉ ◽  
O. RENAUD ◽  
...  
Keyword(s):  
High Resolution ◽  
Three Dimensional ◽  
Living Cells
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