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X-ray microanalysis of hydrated biological specimens
Scanning Electron Microscopy
|January 1, 1982
Summary
This study explores wet scanning electron microscopy (wet-SEM) and X-ray microanalysis for biological soft tissues. The technique successfully identified intracellular calcium in hen oviduct and enzyme activity in rat liver mitochondria.
Area of Science:
- Biological Sciences
- Microscopy
- Biochemistry
Background:
- Traditional scanning electron microscopy (SEM) requires dehydrated and coated specimens, which can alter cellular structures and elemental composition.
- Observing hydrated biological soft tissues under SEM presents challenges due to water content and potential beam damage.
- Developing methods to analyze biological specimens in their native hydrated state is crucial for accurate ultrastructural and elemental analysis.
Purpose of the Study:
- To evaluate the efficacy of wet scanning electron microscopy (wet-SEM) combined with X-ray microanalysis for observing hydrated biological soft tissues.
- To assess the preservation of elemental composition and cytochemical reaction products in unfixed and fixed hydrated specimens.
- To explore the potential clinical applications of this combined technique for surgical specimen analysis.
Main Methods:
- Utilized a wide-angle backscattered electron detector (BED) for low-vacuum SEM (0.3-0.5 torr).
- Examined glutaraldehyde-fixed and unfixed biological soft tissues including hen oviduct, mouse kidney and duodenum, and rat liver mitochondria.
- Correlated backscattered electron images (BEI) with X-ray microanalysis for elemental detection (Ca, P, S, Cl, K).
- Assessed succinate dehydrogenase (SDH-ase) activity in isolated rat liver mitochondria.
Main Results:
- Intracellular secretion granules in fixed hen oviduct preserved calcium (Ca) even after glutaraldehyde fixation.
- X-ray microanalysis detected elements (P, S, Cl, K) in unfixed mouse duodenum and kidney, with hydrated tissues preserving more potassium (K) and less chlorine (Cl) compared to frozen-dehydrated specimens.
- Succinate dehydrogenase (SDH-ase) activity was detectable in hydrated isolated rat liver mitochondria.
Conclusions:
- Wet-SEM combined with X-ray microanalysis offers a valuable tool for assessing localized elements in surgical specimens, providing previously unavailable information.
- Despite limitations in spatial resolution, this technique enables the study of hydrated biological tissues, preserving elemental composition and enzyme activity.
- This method holds promise as a new tool for clinical applications of SEM in analyzing biological samples.