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Le, Dinh To: Analysis, Optimization and Application of AAV Capsid Assembly using Escherichia coli. 2021
Inhalt
Adeno-associated virus capsid protein expression in Escherichia coli and chemically defined capsid assembly
Results
Expression and purification of VP3 proteins.
Figure 1 Expression of VP3wt protein in E.
Assembly and characterization of VLPs.
Cellular uptake of VP3wt VLPs.
Effect of AAP2 on in vitro assembly.
Surface modification but not C-terminal extension is compatible with capsid antibody A20 mAb detection.
Figure 2 Characterization of VP3wt VLPs.
Figure 3 Fluorescent microscopy images of internalization.
Figure 4 Impact of AAP2 on in vitro assembly.
Discussion
Figure 5 The effect of incorporation of a small peptide, either into the 587-loop or at the C-terminus of VP3wt, on in vitro assembly.
Methods
Expression and purification of proteins.
SDS-PAGE and western blot.
In vitro assembly of VLPs.
DLS, AFM Characterization of VLPs.
ELISA.
Cellular uptake of VP3wt VLPs.
Acknowledgements
Introduction
In Vitro Assembly of Virus-Like Particles (VLPs)
Capsid Protein Production
Factors Impacting In Vitro Assembly
pH, Ionic Strength, and Temperature
Nucleic Acids, Scaffolding Protein, and Additives
VLP Characterization
Application of In Vitro-Assembled VLPs
Vaccine Development
Therapeutic Delivery
Conclusions
References