Title

Multimodal production of adeno-associated virus

Document Type

Article

Abstract

Adeno-associated virus (AAV) is an increasingly popular tool in the research laboratory, and use of this viral vector clinically is occurring at an accelerated pace. Nevertheless, despite its popularity, AAV is a relatively cumbersome virus to produce; however, significant efforts have been invested to develop, optimize, and simplify methodology that allows the generation of high-quality AAV with significantly increased production yields. Here we describe multiple modalities for production and purification of AAV particles produced in HEK293 cell cultures using an iodixanol density gradient. We include two methods adapted for harvesting virus from the culture media: Tangential flow filtration (TFF) and polyethylene glycol precipitation (PEGylation). Moreover, we also describe the protocol for anion exchange chromatography, which can be used after the iodixanol gradient as an additional purification step. Last, we provide various protocols for determining virus titer.

Keywords

AAV, AAV production, Digital droplet PCR, Iodixanol gradient, PEGylation, Tangential flow filtration

Publication Date

1-1-2019

Publication Title

Methods in Molecular Biology

ISSN

10643745

Volume

1937

First Page

101

Last Page

124

PubMed ID

30706392

Digital Object Identifier (DOI)

10.1007/978-1-4939-9065-8_6

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