Liposomes freeze fracture electron microscopy

Sternberg, B., Sorgi, F.L., and Huang, L., New structures in complex formation between DNA and cationic liposomes visualized by freeze-fracture electron microscopy, FEBS Letters, 1994, 356, 361-366. 

Phase-separated monolayers and liposomes were characterized by R. Elbert1011 who synthesized saturated and polymerizable fluorocarbon amphiphiles (59, 60, 61) and investigated their mixing behavior with CH2-analogues and natural lipids. In these systems the fluorocarbon compounds are incompatible with hydrocarbon lipids in a wide range of compositions and tend to form domains of pure fluorocarbon and hydrocarbon amphiphiles. The domains can be visualized by freeze-fracture electron microscopy. 

Figure 5.9 Freeze-fracture electron microscopy of phospholipid liposomes imdergoing fusion, after Ellens et al. [66].

Cationic quaternary ammonium compounds such as distearyldimethylammonium-chloride (DSDMAC) used as a softener and as an antistatic, form hydrated particles in a dispersed phase having a similar structure to that of the multilayered liposomes or vesicles of phospholipids 77,79). This liposome-like structure could be made visible by electron microscopy using the freeze-fracture replica technique as shown by Okumura et al. 79). The concentric circles observed should be bimolecular lamellar layers with the sandwiched parts being the entrapped water. In addition, the longest spacings of the small angle X-ray diffraction pattern can be attributed to the inter-lamellar distances. These liposome structures are formed by the hydrated detergent not only in the gel state but also at relatively low concentrations. 

With the freeze-fracture technique, the fracture plane passes through liposomes which are randomly positioned in the frozen sample. Some liposomes will be cut far from their midplane sections, others through their midplane section. Therefore, the analysis of freeze-fracture pictures requires corrections for nonequatorial fracture. Besides, corrections have to be made for the size-dependent probability of a vesicle being in the fracture plane (Jousma et al., 1987 Guiot et al., 1980). Recently, results with a new technique based on electron microscopy was discussed this technique allows analysis not only of liposome size, but also of the number of bilayers (Lauten-schlager et al., 1988). 

Optical stability can be monitored by using electron microscopy, after negative staining for suspensions or after freeze fracturing process. This technique provides indications of the size, shape and number of lamellae of liposomes. 

Morphology of MLs and DLs was evaluated by freeze-fracture and cryotransmission electron microscopy. Both liposomes exhibited single unilamellar vesicles (13). 

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