CHARACTERIZATION OF FROZEN HYDRATED LIPOSOMES BY ELECTRON MICROSCOPY

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Cryo-electron microscopy has been used to characterize the size distribution and shapes of liposomes being developed for delivery of therapeutic cancer drugs. Suitably thin films of the liposomes suspended in aqueous solution were rapidly frozen into liquid ethane. Specimens were cryotransferred into a transmission electron microscope equipped with a post-column energy filter operating at 120 kV accelerating voltage. Energy- filtered images were acquired digitally at low electron dose with a cooled charge couple device detector. It was found that zero- loss imaging provides greatly improved contrast compared with unfiltered imaging because removal of multiply scattered electrons through energy-filtering reduces image blurring caused by chromatic aberration of the objective lens. This technique allowed liposomes to be visualized in vitrified layers that are more than 0.5 micrometers thick. Most of the liposomes were measured to have diameters in the range 20 to 40 nm. However, some could be identified as micelles from their small diameters of approximately 10 nm.