High-Grade Purification of Third-Generation HIV-Based Lentiviral Vectors by Anion Exchange Chromatography for Experimental Gene and Stem Cell Therapy Applications

Methods in Molecular Biology
Hazal Banu OlgunSalih Sanlioglu

Abstract

Lentiviral vectors (LVs) have been increasingly used in clinical gene therapy applications particularly due to their efficient gene transfer ability, lack of interference from preexisting viral immunity, and long-term gene expression they provide. Purity of LVs is essential in in vivo applications, for a high therapeutic benefit with minimum toxicity. Accordingly, laboratory scale production of LVs frequently involves transient cotransfection of 293T cells with packaging and transfer plasmids in the presence of CaPO4. After clearance of the cellular debris by low-speed centrifugation and filtration, lentivectors are usually concentrated by high-speed ultracentrifugation in sucrose cushion. Concentrated viral samples are then purified by anion exchange chromatography (AEX) after benzonase treatment to remove the residual cellular DNA. Here, we describe an improved practical method for LV purification using AEX, useful for experimental studies concerning gene and stem cell therapy.

Citations

Oct 23, 2020·Biotechnology Journal·Ana Sofia MoreiraCristina Peixoto
Nov 2, 2020·Molecular Therapy : the Journal of the American Society of Gene Therapy·Fulya ErendorSalih Sanlioglu
Feb 10, 2021·Signal Transduction and Targeted Therapy·Jote T BulchaGuangping Gao
Feb 13, 2021·Viruses·Christopher Perry, Andrea C M E Rayat
Aug 23, 2021·Advanced Drug Delivery Reviews·Simon StaubachBernd Giebel

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