The majority of adeno-associated virus (AAV) gene therapies are currently developed as frozen formulations (e.g., ≤ - 60 °C) that are challenging to maintain and distribute world-wide. Lyophilization can allow for long-term refrigerated storage and improved shelf-life that lowers long-term cost. Here, we performed a lyophilization feasibility study to assess the ability of several different excipients to stabilize AAV5 during lyophilization and on storage stability. A range of biophysical techniques were used to assess capsid integrity on a molecular level including quantification of externalized DNA, capsid particle size, and capsid monomer percent area. Additionally, transmission electron microscopy was used for the first time to monitor the size and integrity of the capsids subjected to the lyophilization process, and the results supported other characterization methods used in this study. A formulation containing hydroxyectoine and trehalose stabilized capsid structure directly after lyophilization, as observed directly by 5.0 % of internally stained capsids (empty) and indirectly with 7.5 % external DNA. A recombinant human albumin and trehalose formulation stabilized capsid structure on stability as observed by improved external DNA and monomer profiles overtime. Adversely, mannitol crystallization negatively affected capsid structure. Our findings indicate that lyophilization is a viable option to frozen formulation for stabilizing AAV5 drug products.