Monitoring the Stability of Perfluorocarbon Nanoemulsions by Cryo-TEM Image Analysis and Dynamic Light Scattering

Perfluorocarbon nanoemulsions (PFC-NE) are disperse systems consisting of nanoscale liquid perfluorocarbon droplets stabilized by an emulsifier, usually phospholipids. Perfluorocarbons are chemically inert and non-toxic substances that are exhaled after in vivo administration. The manufacture of PFC-NE can be done in large scales by means of high pressure homogenization or microfluidization. Originally investigated as oxygen carriers for cases of severe blood loss, their application nowadays is more focused on using them as marker agents in (19)F Magnetic Resonance Imaging ((19)F MRI). (19)F is scarce in organisms and thus PFC-NE are a promising tool for highly specific and non-invasive imaging of inflammation via (19)F MRI. Neutrophils, monocytes and macrophages phagocytize PFC-NE and subsequently migrate to inflamed tissues. This technique has proven feasibility in numerous disease models in mice, rabbits and mini pigs. The translation to clinical trials in human needs the development of a stable nanoemulsion whose droplet size is well characterized over a long storage time. Usually dynamic light scattering (DLS) is applied as the standard method for determining particle sizes in the nanometer range. Our study uses a second method, analysis of transmission electron microscopy images of cryo-fixed samples (Cryo-TEM), to evaluate stability of PFC-NE in comparison to DLS. Four nanoemulsions of different composition are observed for one year. The results indicate that DLS alone cannot reveal the changes in particle size, but can even mislead to a positive estimation of stability. The combination with Cryo-TEM images gives more insight in the particulate evolution, both techniques supporting one another. The study is one further step in the development of analytical tools for the evaluation of a clinically applicable perfluorooctylbromide nanoemulsion.