Empty/Full Viral Vector Separation - A Hurdle to Clear for Gene Therapies

Adeno-associated viruses (AAVs) are a leading candidate for viral vector delivery of gene therapies. However, AAV manufacturing is time-consuming and challenging due to complex processes, often yielding inconsistent products. Scaling manufacturing of viral vector-mediated gene therapies is hindered by multiple filtration and purification steps, resulting in product loss.

One of the most significant challenges is that inserting payload sequences into AAVs is often inefficient, resulting in empty or partially filled capsids. These capsids are impurities without any therapeutic payload and may produce undesirable immune responses and affect product safety and effectiveness. The ratio of full to empty AAVs is a crucial quality control parameter for obtaining a highly pure and homogeneous population of therapeutic viral vectors. Conventional purification techniques like chromatography are ineffective at separating empty/full capsids and cannot provide batch data on empty/full capsid ratios.

Fortunately, technologies for density-based separation have now made the separations of empty/full viral capsids simpler. A preferred technique is capillary isoelectric focusing, which separates particles based on their isoelectric point. The SCIEX PA800 Plus Pharmaceutical Analysis System offers an optimized workflow for understanding empty/full ratios of AAVs with high resolution. It enables analysis times of less than one hour of analysis per sample, unlike other methods that can take days.

Currently, the gold standard for separating viral capsids is ultracentrifugation. The Optima XPN-series Ultracentrifuge from Beckman Coulter Life Sciences is a serotype-independent ultracentrifugation system that effectively removes impurities like empty and partial capsids during AAV preparation. This system can reduce run times and cut the per-sample cost of purifying AAVs by 250× versus affinity chromatography. Beckman Coulter Life Sciences’ Optima AUC is also a valuable tool for evaluating empty/full ratios in viral preparations, allowing researchers to understand empty/full ratios and quality control parameters for AAV particles.

Viral vectors are crucial to unlocking gene therapy's exciting potential and changing the healthcare landscape. Contact an expert at Danaher Life Sciences to leverage new technologies and accelerate your viral vector preparation workflow.