| 1. |
- 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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| 2. |
- Lundgren, Ted, 1959, et al.
(författare)
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The use of the stable isotope 44Ca in studies of calcium incorporation into dentin.
- 1994
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Ingår i: Journal of microscopy. - 0022-2720. ; 173:Pt 2, s. 149-54
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Tidskriftsartikel (refereegranskat)abstract
- The incorporation into rat incisor dentin of two calcium isotopes, the stable 44Ca and the radioactive 45Ca, was studied using secondary ion mass spectrometry (SIMS) step-scanning and imaging, and autoradiography, respectively. The results demonstrated a time-dependent incorporation of the calcium isotopes into the mineral phase of dentin. With the SIMS step-scanning, detecting 44Ca, the ion yield was high in the odontoblasts 2 min after intravenous injection. After 10 min a marked increase in signal intensity was found at the dentin mineralization front. This result was consistent with those obtained by 45Ca autoradiography; a peak of incorporation occurred 10 min after injection of the isotope. Likewise, localization of 44Ca to the mineralization front could be demonstrated 10 min after injection by SIMS imaging. In images obtained at earlier intervals, no such increase in ion yield could be detected. The results show that the nonradioactive, stable isotope 44Ca can be used as a marker for biomineralization in a similar way to radioactive 45Ca.
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| 3. |
- Trepte, O, et al.
(författare)
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Studies of Porphyrin-containing Specimens Using An Optical Spectrometer Connected To A Confocal Scanning Laser Microscope
- 1994
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Ingår i: Journal of Microscopy. - 0022-2720. ; 176, s. 238-244
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Tidskriftsartikel (refereegranskat)abstract
- A spectrometer has been developed for use with a confocal scanning laser microscope. With this unit, spectral information from a single point or a user-defined region within the microscope specimen can be recorded. A glass prism is used to disperse the spectral components of the recorded light over a linear CCD photodiode array with 256 elements. A regulated cooling unit keeps the detector at 277 K, thereby allowing integration times of up to 60 s. The spectral resolving power, lambda/Delta lambda, ranges from 350 at lambda = 400 nm to 100 at gamma = 700 nm. Since the entrance aperture of the spectrometer has the same size as the detector pinhole used during normal confocal scanning, the three-dimensional spatial resolution is equivalent to that of normal confocal scanning. Light from the specimen is deflected to the spectrometer by a solenoid controlled mirror, allowing fast and easy switching between normal confocal scanning and spectrometer readings. With this equipment, studies of rodent liver specimens containing porphyrins have been made. The subcellular localization is of interest for the mechanisms of photodynamic therapy (PDT) of malignant tumours. Spectroscopic detection is necessary to distinguish the porphyrin signal from other fluorescent components in the specimen. Two different substances were administered to the tissue, Photofrin, a haematoporphyrin derivative (HPD) and delta-amino levulinic acid (ALA), a precursor to protoporphyrin IX and haem in the haem cycle. Both are substances under clinical trials for PDT of malignant tumours. Following administration of these compounds to the tissue, the potent photosensitizer and fluorescent compound Photofrin, or protoporphyrin IX, respectively, is accumulated. For our study Wistar/Furth rats were injected either with Photofrin or with ALA 3-5 h before they were killed. The organs were removed directly after, and snap-frozen in carbon dioxide ice with isopentane. No further staining or fixation procedures were adopted.
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