| 1. |
- Nygren, Håkan, 1952, et al.
(författare)
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Silver deposition on freeze-dried cells allows subcellular localization of cholesterol with imaging TOF-SIMS.
- 2004
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Ingår i: Journal of microscopy. - : Wiley. - 0022-2720 .- 1365-2818. ; 215:Pt 2, s. 156-61
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Tidskriftsartikel (refereegranskat)abstract
- Imaging time-of-flight secondary ion mass spectrometry (TOF-SIMS) was used for characterization and subcellular localization of organic ions in leucocytes adhering to glass surfaces. The cells were fixed by freeze drying in 0.15 m ammonium formate buffer at pH 7.2-7.4. The freeze-dried cells were sputter-coated with silver, and the silver surface was analysed with imaging TOF-SIMS. TOF-SIMS spectra were recorded by scanning the primary ion beam over the analysis area and acquiring positive mass spectra of the ions leaving the surface. The relative brightness of each pixel within the analysis area reflects the signal intensity of a selected ion in that pixel. Data were collected separately at high mass resolution m/delta m > 7000 and at high lateral resolution (= 0.5 micro m). The images were analysed by principal component analysis (PCA). The glass-adhering cells showed a well defined attachment area with a diameter of up to 20 micro m, and an equally well defined cell body, containing the nucleus, with a diameter of 8-10 micro m. On the raw data images, the obtained cholesterol distributions were consistent with a higher cholesterol content of the cell membrane in the attachment area than in the cell body. Using PCA analysis, silver-cationized molecular cholesterol was found localized mainly in the attachment area of the cells. Cholesterol was also seen at higher concentration in circular spots of
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| 2. |
- Johansson, Göran A., et al.
(författare)
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Exploring the use of soft X-ray microscopy for imaging subcellular structures of the inner ear
- 2004
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Ingår i: Journal of Microscopy. - : Wiley. - 0022-2720 .- 1365-2818. ; 215, s. 203-212
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Tidskriftsartikel (refereegranskat)abstract
- The soft X-ray microscope at the Lawrence Berkeley National Laboratory was developed for visualization of biological tissue. Soft X-ray microscopy provides high-resolution visualization of hydrated, non-embedded and non-sectioned cells and is thus potentially an alternative to transmission electron microscopy. Here we show for the first time soft X-ray micrographs of structures isolated from the guinea-pig inner ear. Sensory outer hair cells and supporting pillar cells are readily visualized. In the hair cells, individual stereocilia can easily be identified within the apical hair bundle. The underlying cuticular plate is, however, too densely composed or too thick to be clearly visualized, and thus appears very dark. The cytoplasmic structures protruding from the cuticular plates as well as the fibrillar material surrounding and projecting from the cell nuclei can be seen. In the pillar cells the images reveal individual microtubule bundles. Soft X-ray images of the acellular tectorial membrane and thin two-layered Reissner's membrane display a level of resolution comparable to low-power electron microscopy.
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| 3. |
- Mannelquist, Anders, et al.
(författare)
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Near field optical microscopy in aqueous solution : implementation and characterization of a vibrating probe
- 2002
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Ingår i: Journal of Microscopy. - : Wiley. - 0022-2720 .- 1365-2818. ; 205:1, s. 53-60
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Tidskriftsartikel (refereegranskat)abstract
- Near field optical microscopy (NSOM) is one of the possible solutions to circumvent the diffraction limit, but the control of the optical probe in solution has been a technical challenge for practical applications. Most recently, it has been shown that the pipette used in the scanning ion conductance microscope can be modified to form a high resolution near field optical probe. When combined with a novel distance modulation mechanism, a robust near field microscope can be constructed for operation in aqueous solution. In this paper, we present technical details of this design and a further characterization of the NSOM system for imaging in solution. Fundamental limitations of this approach in comparison to other systems are also discussed. Based on the current technology, it is concluded that better than 50 nm resolution should be achievable with this technique for fluorescence, as well as fluorescence resonance energy transfer, imaging of biological specimens.
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| 4. |
- Wählby, Carolina, et al.
(författare)
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Combining intensity, edge, and shape information for 2D and 3D segmentation of cell nuclei in tissue sections
- 2004
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Ingår i: Journal of Microscopy. - : Wiley. - 0022-2720 .- 1365-2818. ; 215:1, s. 67-76
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Tidskriftsartikel (refereegranskat)abstract
- We present a region-based segmentation method in which seeds representing both object and background pixels are created by combining morphological filtering of both the original image and the gradient magnitude of the image. The seeds are then used as starting points for watershed segmentation of the gradient magnitude image. The fully automatic seeding is done in a generous fashion, so that at least one seed will be set in each foreground object. If more than one seed is placed in a single object, the watershed segmentation will lead to an initial over-segmentation, i.e. a boundary is created where there is no strong edge. Thus, the result of the initial segmentation is further refined by merging based on the gradient magnitude along the boundary separating neighbouring objects. This step also makes it easy to remove objects with poor contrast. As a final step, clusters of nuclei are separated, based on the shape of the cluster. The number of input parameters to the full segmentation procedure is only five. These parameters can be set manually using a test image and thereafter be used on a large number of images created under similar imaging conditions. This automated system was verified by comparison with manual counts from the same image fields. About 90% correct segmentation was achieved for two- as well as three-dimensional images.
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| 5. |
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| 6. |
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| 7. |
- Adler, Jeremy
(författare)
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The unitary scale bar : human and machine readable
- 2008
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Ingår i: Journal of Microscopy. - : Wiley. - 0022-2720 .- 1365-2818. ; 230:1, s. 163-166
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Tidskriftsartikel (refereegranskat)abstract
- A format is described for a scale bar that encodes the length represented within the structure of the bar itself, thereby removing the need for any supporting text. Although the 'unitary' scale bar has a conventional appearance it is also machine readable and therefore retains information about the scale even when the file format is changed. The format is based on the metre and is suitable for all terrestrial applications.
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| 8. |
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| 9. |
- Almqvist, Nils, et al.
(författare)
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Micromechanical and structural properties of a pennate diatom investigated by atomic force microscopy
- 2001
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Ingår i: Journal of Microscopy. - : Wiley. - 0022-2720 .- 1365-2818. ; 202:3, s. 518-532
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Tidskriftsartikel (refereegranskat)abstract
- The mechanisms behind natural nanofabrication of highly structured silicas are increasingly being investigated. We have explored the use of a standard Nanoscope III Multimode atomic force microscope (AFM) to study the silica shell of diatoms. The delicate structures of the shell surface of the diatom Navicula pelliculosa (Breb.) Hilse were imaged and the shell's micromechanical properties were measured semi-quantitatively with a resolution down to approximately 10 nm. The technique to measure elasticity and hardness with the AFM was demonstrated to be useable even on these hard glass-like surfaces, Different experimental configurations and evaluation methods were tested, They gave a consistent result of the shell micromechanical properties, The first results showed that the diatom shell's overall hardness and elasticity was similar to that of known silicas. However, regions with different mechanical proper ties were distinguished. The elastic modulus varied from 7 to 20 GPa, from 20 to 100 GPa and from 30 to hundreds of GPa depending on the location. In general, the hardness measurements showed similar spatial differences, The hardness values ranged from 1 to 12 GPa but one specific part of the shell was even harder. Hence, certain localized regions of the shell were significantly harder or more elastic. These regions coincide with known characteristic features and mechanisms appearing at the different stages of the shell's growth. These results show that this method serves as a complementary tool in the study of silica biomineralization, and can detect eventual crystalline phases.
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| 10. |
- Andersson-Engels, Stefan, et al.
(författare)
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Time-resolved and Wavelength-resolved Spectroscopy In 2-photon-excited Fluorescence Microscopy
- 1994
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Ingår i: Journal of Microscopy. - 0022-2720. ; 176, s. 195-203
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Tidskriftsartikel (refereegranskat)abstract
- Two-photon excited fluorescence spectroscopy has been performed at a microscopic scale in combination with normal, white-light microscopy. This gave simultaneously a spectral resolution of 20 nm and a temporal resolution of 20 ps, from a volume element less than 5 mu m in all three dimensions. The sample was excited with the light from a continuously mode-locked Ti:sapphire laser that was focused on the sample in a fluorescence microscope. A polychromator and a streak-camera were used for detection. The method has been used on tissue, plant and paper samples. It has also been demonstrated how substances naturally occurring in the samples can be identified from their spectroscopic properties and the spatial distribution of these substances can be observed.
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