Cochleate characterization by cryogenic electron microscopy methods: Cryo-TEM and Cryo-SEM

Cochleates are lipid microstructures consisting of negatively charged phospholipid bilayers rolled up into cigar-like spirals, which are formed through the interactions with multivalent counter-ions used as bridging agents. This lipid system was shown to have potential as a drug carrier, primarily for oral delivery. In this study, we describe the structure of cochleates made of dioleoylphosphatidylserine (DOPS), a negatively charged lipid, and calcium that induce the bridging of lipid layers and complex formation. By using advanced cryogenic electron microscopy (CryoEM) methods, cryo-TEM and cryo-SEM, we are able to explore local and global structural details of the cochleates at length scales ranging from the nano to the micro-scale, at high resolution, and at conditions relating to the hydrated, native state. We reveal the nano-layers of cochleates and provide evidence to the mechanism of their formation, and show that that full cochleates and hollow ones can be created, as well as cone-shaped structures. Also, cochleates may be formed by the spiraling of more than one bilayer lamella. The structural insights from this study shed light on the complexation process, which ultimately can impact the delivery of drugs by such carriers. (C) 2015 Elsevier B.V. All rights reserved.