Lentiviral Vector Purification Using Genetically Encoded Biotin Mimic in Packaging Cell

Area of Science:

• Biotechnology
• Molecular Biology
• Gene Therapy

Background:

• Lentiviral vectors (LVs) are increasingly used in research and clinical settings.
• Current LV purification methods are inefficient, costly, and yield low recovery rates, hindering widespread application.

Purpose of the Study:

• To develop a streamlined, one-step purification process for lentiviral vectors (LVs).
• To enhance LV purity and yield using a novel biotin-streptavidin-based system.

Main Methods:

• Genetically engineered HEK293T cells to display a biotin-mimicking peptide (cTag8) on LVs during budding.
• Utilized streptavidin magnetic beads for LV capture via cTag8.
• Employed competitive elution with biotin to release purified LVs.

Main Results:

• Achieved overall LV yields of ≥60% with high purity.
• Demonstrated >3-log reduction in DNA and >2-log reduction in host cell proteins.
• Preliminary scalability testing using monolith affinity chromatography showed a 20% yield.

Conclusions:

• The developed one-step purification method offers high yield and purity for LVs.
• This approach effectively removes contaminants and is potentially scalable for clinical applications.
• The cTag8 system provides a valuable tool for advancing LV research and therapeutic use.