Heptane and isooctane as embedding fluids for high-pressure freezing of Petunia ovules followed by freeze-substitution
| Autor: | A.C. van Aelst, M.H. Thijssen, J. L. Van Went |
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| Jazyk: | angličtina |
| Rok vydání: | 1998 |
| Předmět: |
Laboratorium voor Plantencelbiologie
Histology Petunia Pathology and Forensic Medicine Cryofixation chemistry.chemical_compound Petunia x hybrida Congelation Acetone Ovule Heptane High-pressure freezing Chromatography biology biology.organism_classification 1-hexadecene Laboratory of Plant Cell Biology Isooctane chemistry Freeze substitution Melting point Ultrastructure EPS Freeze-substitution |
| Zdroj: | Journal of Microscopy 192 (1998) Journal of Microscopy, 192, 228-235 |
| ISSN: | 0022-2720 |
| Popis: | In comparison with other fixation methods, high-pressure freezing and freeze-substitution of Petunia ovules lead to improved ultrastructural preservation of all tissues. Crucial for adequate high-pressure freezing is the absence of air in the specimen sandwich; air has to be replaced by an embedding fluid. Frequently, 1-hexadecene is used for this purpose. Using 1-hexadecene as an embedding fluid resulted in only 5–10% of Petunia ovules being preserved without disturbance of the ultrastructure due to ice-crystal damage. Since 1-hexadecene is not soluble in acetone at − 90 °C, freeze-substitution is hindered when ovules remained completely surrounded by it; this results in recrystallization when the temperature is raised. We tested and compared the suitability of heptane and isooctane as embedding fluids for high-pressure freezing and freeze-substitution, reasoning that because of their low melting points and low relative densities, phase separation during freeze-substitution would result in complete exposure of the ovules to the substitution medium, leading to adequate freeze-substitution. Using either heptane or isooctane as an embedding fluid yielded up to 90% ice-crystal-free ovules. Both compounds, however, have some damaging effects on the outer one or two cell layers of the ovule, but not on the inner tissues. |
| Databáze: | OpenAIRE |
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