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High-pressure freezing causes structural alterations in phospholipid model membranes
K Semmler1, J Wunderlich, W Richter
1Institute of Ultrastructure Research, Friedrich-Schiller-University Jena, Germany.
Journal of Microscopy
|July 23, 1998
Summary
High-pressure freezing (HPF) alters the lipid arrangement in mixed phospholipid membranes, changing their macroripple-phase morphology. Pure dipalmitoyl-phosphatidylcholine (DPPC) membranes remain unaffected by HPF, suggesting pressure-induced relaxation in binary systems.
Area of Science:
- Biophysics
- Materials Science
Background:
- Phospholipid model membranes are crucial for understanding biological systems.
- High-pressure freezing (HPF) is a technique used to preserve sample structure for microscopy.
- Lipid packing and phase behavior are fundamental properties of membranes.
Purpose of the Study:
- To investigate the effect of high-pressure freezing (HPF) on the lipid arrangement in phospholipid model membranes.
- To analyze morphological changes in liposomes composed of dipalmitoyl-phosphatidylcholine (DPPC) and a branched-chain phosphocholine under HPF.
- To evaluate the performance of a newly designed sample holder for HPF.
Main Methods:
- Liposomes of pure DPPC and DPPC mixed with a branched-chain phosphocholine were prepared.
- Samples were frozen using conventional plunging into liquid propane and high-pressure freezing (HPF).
- Freeze-fracture electron microscopy was employed to analyze the lipid arrangement and morphology.
- A novel sandwiched copper platelet sample holder for the Balzers HPM010 HPF machine was utilized and tested.
Main Results:
- HPF significantly altered the macroripple-phase morphology in the two-component DPPC liposome system.
- Pure DPPC bilayer membranes showed no observable alterations after HPF.
- Control experiments confirmed that ethanol, used as a pressure transfer medium, did not cause the observed changes.
- The newly designed sandwich sample holder allowed for HPF of thin samples (20-100 microns) and provided temperature control.
- Despite mass reduction, the new holder did not improve HPF cooling rates (4000 °C/s), indicating cryogen velocity is the limiting factor.
Conclusions:
- HPF induces structural modifications in mixed phospholipid membranes, likely due to pressure-induced relaxation of unstable lipid packing.
- Pure DPPC membranes are more stable under HPF conditions.
- The developed sandwich sample holder is effective for HPF of fluid samples, though cooling rates are primarily determined by the cryogen.
- Cryogen velocity is the main determinant of cooling rates in high-pressure freezing.