Achieve significantly increased adenovirus yield with density gradient ultracentrifugation: a comparative study

September 13, 2022


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Adenoviral (AdV) vectors are highly effective tools for gene therapy (GT), oncolytic virotherapy, and vaccines due to their high gene transduction efficiencies, safety, and tunability. Although numerous technologies exist for the downstream purification of AdV, several critical manufacturing challenges still exist -- including scalability, low recovery, and the removal of highly similar impurities such as empty capsids.

Two of the most prominent AdV purification techniques include density gradient ultracentrifugation (DGUC) and ion-exchange chromatography (IEC). Although IEC is well established as a scalable approach to biomolecule purification with short separation times, it suffers from several limitations in the context of AdV, including low binding capacity, serotype dependency, and a tradeoff between yield and purity. DGUC, however, is a robust, serotype-independent method that offers improved product recovery by exploiting subtle differences in the buoyant density between full, therapeutically active AdV particles and other process impurities.

Here, we directly compare both techniques and demonstrate significant improvements in product critical quality attributes (CQAs) including >200-fold increases in yield and concentration for the DGUC-purified AdV5. In addition, we will discuss approaches to maximize throughput and efficiency when scaling up or down, which are universally applicable across a range of therapeutic platforms such as AdV, adeno-associated virus (AAV), and more.

Watch this webinar to learn: 

  • How DGUC and IEC perform with respect to various AdV CQAs
  • Key considerations for choosing DGUC or IEC for AdV purification
  • Solutions for rapid implementation, optimization, and scaling of DGUC for purification 

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Speaker Bio

spINSIGHT Ultracentrifugation is preferred for high purity EV isolation Shawn SternishaShawn Sternisha, Ph.D.
Senior Field Applications Scientist, Beckman Coulter Life Sciences

Shawn Sternisha is currently a Senior Field Applications Scientist in the Biotechnology Business Unit at Beckman Coulter Life Sciences where he is primarily focused on the development of novel preparative and analytical ultracentrifuge applications. Shawn also supports market activities across all centrifugation product lines and is responsible for KOL and partnership management. Shawn attended Illinois State University for his undergraduate studies and went on to earn his Ph.D. in Biochemistry at Florida State University. Shawn is broadly interested in cell and gene therapy, extracellular vesicles, enzyme structure-function relationships, and expanding applications of ultracentrifugation. Shawn currently resides in Miami, FL with his wife Patty and their dog Ruby. In his free time, he enjoys cooking, running, traveling, and learning Spanish.