Ion-Exchange Resins
At present, the research products based on recombinant adeno-associated virus (rAAV) are facing the challenge of clinical development. Even high-purity preparations contain impurities related to process and product identification, characterization and batch-by-batch control. Creative Biolabs focuses on the development of viral vectors, and has proposed the chromatographic purification scheme for rAAV, including ion exchange chromatographic (IEC), to obtain viruses with high yield, purity and bioavailability.
Introduction of IEC for AAV Vector Purification
rAAV has been proved to be an important potential DNA delivery vector. In vivo gene transduction based on AAV vectors relies on laborious procedures for producing high titer vectors. In some rAAV vector production schemes, the contamination of auxiliary viruses is an important safety issue in clinical gene delivery. In order to effectively remove impurities such as host cell proteins and empty vector particles, purification steps are needed to meet the final product specifications. At present, a large number of studies have shown that rAAV can be purified by ion exchange resin based on the charge properties of the protein exposed to the viral capsid.
IEC Development for AAV Vector Purification at Creative Biolabs
IEC allows the development of general purification schemes for different AAV serotypes. The net charge of protein on the AAV surface depends on the acidity and basicity of the exposed amino acid group. Therefore, salt concentration and pH regulation are necessary to optimize the binding of virus particles to chromatographic resins. At present, Creative Biolabs has successfully used a cation-exchange SP-Sepharose-HP column and an anion-exchange source 15Q column to separate and purify AAV serotypes 2 and 5.
In addition, IEC has great potential in separating AAV empty vector particles. Among many AAV vector preparations, a prominent pollutant is the lack of genomic vector particles, which can significantly increase the dose of AAV capsid protein and lead to unnecessary immune consequences. We have successfully separated AAV-8 empty vector and full vector particles by shallow gradient and strong anion exchange chromatography, which also provides a new idea for their quantitative analysis.
Figure 1. An example of the purification protocol for AAV. (Potter, 2014)
Features of Our Services
- Provide rapid and simple concentration and purification of AAV vector services, regardless of the source of these raw materials from cell lysates or cell culture media.
- Customized purification protocols based on different serotype vectors, most of the contaminated cell fragments are removed, while the contaminated DNA and RNA are reduced.
- Purified AAV vector has high activity in in vitro transduction experiments.
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Reference
- Potter, M.; et al. (2014). A simplified purification protocol for recombinant adeno-associated virus vectors. Molecular Therapy-Methods & Clinical Development. 1: 14034.
