Chapter 28
Application of Cryogenic Transmission Electron Microscopy
for Evaluation of Vaccine Delivery Carriers
Hui Qian, Yimei Jia, and Michael J. McCluskie
Abstract
Cryogenic transmission electron microscopy (Cryo-TEM) enables visualizing the physicochemical structure of nanocarriers in solution. Here, we demonstrate the typical applications of Cryo-TEM in characterizing archaeosome-based vesicles as antigen carriers, including the morphology and size of vaccine carriers.
Cryo-TEM tomography, incorporated with immunogold labeling for identifying and localizing the antigens, reveals the antigen distribution within archaeosomes in three dimensions (3D).
Key words Cryo-TEM, Plunge freezing, Electron Tomography, Immunogold labeling, Vaccine
delivery carriers
1 Introduction
Cryofixation not only preserves the biomolecular structure near to
its native state but also enables the structural study in a transmission
electron microscope (TEM) for biological materials in aqueous
solution [1]. Along with the advanced EM instrumentation and
computing technology development, nowadays the structural resolution of biomolecules is revealed down to sub-A ˚ using cryogenic
TEM (Cryo-TEM) single particle analysis (SPA) [2, 3]. Cryo-TEM
has also been widely used for characterization of drug delivery
nanocarriers, such as inorganic colloid nanoparticles, lipid-based
nanoparticles, or virus-like nanoparticles (VLP) (Fig. 1) [4]. Since
the physicochemical characteristics, including size, shape, plasticity,
and complex nanostructure, play a significant role in vaccine delivery effectiveness [5], each procedure of Cryo-TEM (Fig. 2) is
critical to properly characterize nanocarriers.
In general, after the production of nanocarriers, the samples
need to be optimized suitable for TEM analysis, such as the nanoparticle concentration. Sometimes preparation may involve further
treatment for nanocarriers. For instance, nanocarriers are negatively
stained with heavy metals to enhance image contrast; buffer is
Blaine A. Pfeifer and Andrew Hill (eds.), Vaccine Delivery Technology: Methods and Protocols, Methods in Molecular Biology,
vol. 2183, https://doi.org/10.1007/978-1-0716-0795-4_28, © Springer Science+Business Media, LLC, part of Springer Nature 2021
499
Application of Cryogenic Transmission Electron Microscopy
for Evaluation of Vaccine Delivery Carriers
Hui Qian, Yimei Jia, and Michael J. McCluskie
Abstract
Cryogenic transmission electron microscopy (Cryo-TEM) enables visualizing the physicochemical structure of nanocarriers in solution. Here, we demonstrate the typical applications of Cryo-TEM in characterizing archaeosome-based vesicles as antigen carriers, including the morphology and size of vaccine carriers.
Cryo-TEM tomography, incorporated with immunogold labeling for identifying and localizing the antigens, reveals the antigen distribution within archaeosomes in three dimensions (3D).
Key words Cryo-TEM, Plunge freezing, Electron Tomography, Immunogold labeling, Vaccine
delivery carriers
1 Introduction
Cryofixation not only preserves the biomolecular structure near to
its native state but also enables the structural study in a transmission
electron microscope (TEM) for biological materials in aqueous
solution [1]. Along with the advanced EM instrumentation and
computing technology development, nowadays the structural resolution of biomolecules is revealed down to sub-A ˚ using cryogenic
TEM (Cryo-TEM) single particle analysis (SPA) [2, 3]. Cryo-TEM
has also been widely used for characterization of drug delivery
nanocarriers, such as inorganic colloid nanoparticles, lipid-based
nanoparticles, or virus-like nanoparticles (VLP) (Fig. 1) [4]. Since
the physicochemical characteristics, including size, shape, plasticity,
and complex nanostructure, play a significant role in vaccine delivery effectiveness [5], each procedure of Cryo-TEM (Fig. 2) is
critical to properly characterize nanocarriers.
In general, after the production of nanocarriers, the samples
need to be optimized suitable for TEM analysis, such as the nanoparticle concentration. Sometimes preparation may involve further
treatment for nanocarriers. For instance, nanocarriers are negatively
stained with heavy metals to enhance image contrast; buffer is
Blaine A. Pfeifer and Andrew Hill (eds.), Vaccine Delivery Technology: Methods and Protocols, Methods in Molecular Biology,
vol. 2183, https://doi.org/10.1007/978-1-0716-0795-4_28, © Springer Science+Business Media, LLC, part of Springer Nature 2021
499