Enhancing sanitization for AVB Sepharose resin in AAV vector purification

Abstract

Affinity chromatography is a critical step in gene therapy for capturing adeno-associated virus (AAV) vectors. However, the high cost of affinity resin needs effective cleaning and sanitization strategies to enable multi-cycle usage. This study evaluated various combinations of cleaning reagents, including alcohol, low concentrations of sodium hydroxide (NaOH), and acids, against a broad range of microbial strains, particularly acid- and alkaline-resistant species, to identify enhanced sanitization protocols for both pre-use and post-use. A solution comprising 100 mM acetic acid with 2 % benzyl alcohol, alongside 10 mM NaOH with 2 % benzyl alcohol, was identified as effective. This new cleaning and sanitization strategy, incorporating both pre- and post-use cleaning, was successfully implemented, allowing for up to six column cycles without product carryover between cycles. Notably, this strategy does not compromise process yield or product quality. It provides effective microbial control during AAV purification using AVB Sepharose resin, while also preserving resin integrity, reducing the risk of microbial contamination and product carryover, lowering AAV manufacturing costs, and ultimately enhancing the quality and reliability of gene therapy product manufacturing.