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Liposomes, Exosomes, and Virosomes: From Modeling Complex
Membrane Processes to Medical Diagnostics and Drug Delivery
Thursday Speaker Abstracts
43
Monitoring Extracellular-Vesicles Dynamics at the Nanoscale by Liquid-Cell TEM
Max Piffoux
1
, Amanda Brun
1
, Florence Gazeau
1
, Damien Alloyeau
2
.
1
Laboratoire matière et systèmes complexes, Paris, France,
2
Laboratoire matériaux et
phénomènes quantiques, Paris, France.
Introduction:
Exosomes and microvesicles are promising biotherapies that could potentially replace
conventional cell therapies. On the road toward the routine use of these nano-objects in clinics,
the nanoscale characterization of these complex soft materials in liquid environment has to be
improved. Transmission electron microscopy has been a method of choice to image
microvesicles embedded in amorphous ice. Nevertheless the effects of freezing processes on the
membrane remains unclear and make impossible dynamic observations. Here we describe the
use of liquid-cell transmission electron microscopy (LCTEM) for the dynamic characterization
of extracellular vesicles (EV) in PBS media.
Results:
LCTEM consists in imaging the dynamics of nano-objects in an encapsulated liquid solution
within an electron-transparent microfabricated cell. We demonstrate that this recent in situ
technique allows the observation of EV in their native state without any prior staining and
provides the unique opportunity to explore their behavior and structural characteristics in real
time with nanometer resolution. We determined relevant parameters for EV based therapy, such
as their size distribution, concentration, phosphatidyl-serine content (using gold labeled annexin
V) and magnetic nanoparticle loading. Besides, the morphological analyses of EVs in liquid,
through the measurement of their reduced volume, allow studying their size-dependent physical
properties. Using real-time imaging (up to 25 frames/s), EV motion and dynamics in liquid is
analyzed. We reveal that their Brownian motion is slowed by a 100 fold, due to
electrostatic/covalent interactions with the membranes of the liquid-cell. Remarkably, the
detection of fusion events shows that LCTEM could also open up a new way to study membrane
dynamics. Further works are in progress to investigate sample purity and other dynamical
processes, such as EV labeling or osmotic shocks, by controlling the composition of the media.
Lipoproteins in Hedgehog Release and Signaling
Suzanne Eaton
Max Planck Institute for Biophysical Chemistry, Goettingen, Germany
No Abstract