Imaging of small particles using wide-field confocal microscopy

2003 ◽  
Author(s):  
Stephen P. Morgan ◽  
N. B. E. Sawyer ◽  
Michael G. Somekh ◽  
Chung Wah See ◽  
B. Y. Shekunov ◽  
...  
Keyword(s):  
Confocal Microscopy ◽  
Wide Field ◽  
Small Particles

Wide field, phase measuring confocal microscopy of small particles

2003 ◽  
Author(s):  
N. B. E. Sawyer ◽  
Stephen P. Morgan ◽  
Michael G. Somekh ◽  
Chung Wah See ◽  
B. Y. Shekunov ◽  
...  
Keyword(s):  
Confocal Microscopy ◽  
Wide Field ◽  
Small Particles ◽  
Field Phase

scholarly journals Dermoscopy-guided reflectance confocal microscopy of skin using high-NA objective lens with integrated wide-field color camera

2016 ◽  
Author(s):  
David L. Dickensheets ◽  
Seth Kreitinger ◽  
Gary Peterson ◽  
Michael Heger ◽  
Milind Rajadhyaksha
Keyword(s):  
Confocal Microscopy ◽  
Reflectance Confocal Microscopy ◽  
Objective Lens ◽  
Wide Field

Fast Imaging of Single Molecules and Nanoparticles by Wide-Field Microscopy and Spectrally Resolved Confocal Microscopy

2000 ◽  
Vol 1 (4) ◽  
pp. 291-298 ◽  
Author(s):  
W.G.J.H.M. van Sark ◽  
P.L.T.M. Frederix ◽  
D.J. van den Heuvel ◽  
M.A.H. Asselbergs ◽  
I. Senf ◽  
...  
Keyword(s):  
Confocal Microscopy ◽  
Single Molecules ◽  
Fast Imaging ◽  
Wide Field ◽  
Spectrally Resolved

Reliability of Hoechst 33342 Staining under Wide-Field Microscopy for Evaluation of the Nuclear Status of Living Dog Oocytes

2012 ◽  
Vol 18 (3) ◽  
pp. 483-492 ◽  
Author(s):  
Martine Chebrout ◽  
Pierre-Gaël Adenot ◽  
Karine Reynaud ◽  
Sylvie Chastant-Maillard
Keyword(s):  
Confocal Microscopy ◽  
Wide Field ◽  
Hoechst 33342 ◽  
Developmental Potential ◽  
Dna Staining ◽  
Hoechst Staining ◽  
Metaphase Ii Oocytes

AbstractDue to the marked cytoplasmic opacity of canine oocytes, the diagnosis of their nuclear status is difficult. The objective of the present study was to evaluate the accuracy of Hoechst staining observed under epifluorescence wide-field microscopy [living oocyte observation (LivOO)] by comparison to a reference technique [DNA staining with ethidium homodimer-2 under confocal microscopy; fixed oocyte observation (FixOO)]. Four Hoechst 33342 concentrations (200 ng, 500 ng, 1 μg, 2 μg/mL) were tested and 1 μg/mL was the lowest one with the lowest proportion of oocytes in which DNA was missed. At this concentration, LivOO procedure did not affect the degeneration rate. On 379 oocytes observed individually with the two techniques successively, diagnosis of meiosis resumption by LivOO was exact in 87.3% of the cases, but the meiosis resumption rate was underestimated (23.5% versus 34.3% with FixOO; p < 0.001). Diagnosis for metaphase II was exact in 80% of the cases, but LivOO detected only 72.7% of the metaphase II oocytes present. Metaphase rates did not differ between LivOO and FixOO. This study contributes to a better interpretation of in vitro maturation results. The developmental potential of metaphase II canine oocytes sorted after Hoechst staining is to be evaluated.

Video-Confocal Microscopy: An “Electronic-Pinhole” Method using Narrow-Field Illumination and Wide-Field Image Detection.

1999 ◽  
Vol 5 (S2) ◽  
pp. 470-471
Author(s):  
Pier A. Benedetti ◽  
Valter Evangelista ◽  
Dante Guidarini ◽  
Stefano Vestri
Keyword(s):  
Confocal Microscopy ◽  
Optical Microscopy ◽  
State Of The Art ◽  
Single Point ◽  
Three Dimensional ◽  
Axial Direction ◽  
Wide Field ◽  
Biological Investigation ◽  
Photo Damage ◽  
Set Up

State of the art in optical microscopyOptical microscopy is still and increasingly one of the most valuable tools in biological investigation. In particular, confocal microscopies are capable of achieving best performances in the study of three-dimensional fluorescent and reflecting specimens. Nevertheless, current techniques adopted in confocal microscopy present some drawbacks and limitations that stimulate to devise and set-up further techniques, suited to a wider range of applications.Advantages of confocal microscopes mainly correspond to an improved spatial resolution, especially in the axial direction. Depending on the narrow-field scanning approach used, there are two main forms of confocal microscopes: single-point (SP) and multi-point (MP) ones. Unfortunately, SP confocal microscopes require the use of lasers as illumination sources with consequent high costs and scarce spectral flexibility. Moreover, specimen photo-damage due to relatively high instantaneous irradiation doses involved, can often limit their investigative capabilities. On the other hand, proposed MP confocal microscopes still rely on the revolving-disk approach and exhibit a relatively low luminous efficiency, substantial constructional complexity, and limited contrast in the study of thick fluorescent objects.

Confocal microscopy, image restoration, and nuclear structure

1993 ◽  
Vol 51 ◽  
pp. 146-147
Author(s):  
Peter Shaw ◽  
Alison Beven ◽  
David Rawlins ◽  
Martin Highett
Keyword(s):  
Confocal Microscopy ◽  
Functional Organization ◽  
Confocal Imaging ◽  
Rrna Genes ◽  
Wide Field ◽  
Image Deconvolution ◽  
Microscopy Image ◽  
Confocal Images ◽  
Confocal Microscopy Image

Fluorescent in situ hybridization and 3-D confocal microscopy have been used to study the arrangement of genes and transcripts in interphase plant nuclei. Wide-field fluorescence microscopy coupled with computer deconvolution has also been used on the same specimens and gives comparable results to confocal imaging. When image deconvolution is applied to confocal images, these show a significant improvement. A comparison of confocal and wide-field (CCD) images of the same specimen (a plant nucleolus labelled with probe to the rRNA genes) is shown in Figure 1. The deconvoluted wide-field data (Fig 1b) and the raw confocal data (Fig 1c) are remarkably similar; the former is somewhat clearer. The deconvoluted confocal data (Fig 1d) is clearest of all.Double-stranded DNA and single-stranded RNA probes to the 45S and 5S RNA genes have been used to investigate the functional organization of the nucleolus in pea root tissue. In situ hybridization was carried out on vibratome slices approximately 50μm thick (about 3 cells).

scholarly journals Wide-field imaging combined with confocal microscopy using a miniature f/5 camera integrated within a high NA objective lens

2017 ◽  
Vol 42 (7) ◽  
pp. 1241 ◽  
Author(s):  
David L. Dickensheets ◽  
Seth Kreitinger ◽  
Gary Peterson ◽  
Michael Heger ◽  
Milind Rajadhyaksha
Keyword(s):  
Confocal Microscopy ◽  
Objective Lens ◽  
Wide Field ◽  
Field Imaging ◽  
Wide Field Imaging

scholarly journals Confocal microscopy on the beamline: novel three-dimensional imaging and sample positioning

2012 ◽  
Vol 45 (5) ◽  
pp. 936-943 ◽  
Author(s):  
I. Khan ◽  
R. Gillilan ◽  
I. Kriksunov ◽  
R. Williams ◽  
W. R. Zipfel ◽  
...  
Keyword(s):  
Confocal Microscopy ◽  
Second Harmonic ◽  
Imaging Techniques ◽  
Three Dimensional ◽  
Protein Crystal ◽  
Protein Crystals ◽  
Wide Field ◽  
Three Dimensional Imaging ◽  
Biological Studies ◽  
Confocal Optics

Confocal microscopy, a technique that has been extensively applied in cellular biological studies, may also be applied to the visualization and three-dimensional imaging of protein crystals at high resolution on synchrotron beamlines. Protein crystal samples are examined using a commercially available confocal microscope adapted for cryogenic use. A preliminary test using a custom confocal design adapted for beamline use is also presented. The confocal optics configuration is compatible with nonlinear imaging techniques such as two-photon excited fluorescence imaging and second harmonic generation. The possibilities of this method are explored using two modes: fluorescence and reflection confocal. In fluorescence mode, small amounts of dye are introduced into the crystal through soaking or growth conditions. Under such conditions, protein crystals are easily resolved from salts and amorphous precipitates, which do not generally take up dye. Reflection mode, which does not require dye, still exhibits greater resolution and sensitivity to surface detail than conventional wide-field microscopy as a result of the confocal optics configuration. The inherent three-dimensional nature of the method means that on-axis sample views (along the direction of the X-ray beam) can be reconstructed from an off-axis configuration, simplifying the beamline setup and providing uniquely detailed views of cryogenically cooled crystals.

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