Classification and quantification of full vs. empty adeno-associated virus reference capsids via infrared attenuated...

Area of Science:

• Biotechnology
• Analytical Chemistry
• Molecular Biology

Background:

• Adeno-associated virus (AAV) vectors are crucial for gene therapy, offering stability and long-term gene expression.
• Current AAV analysis methods are complex, costly, and unsuitable for real-time manufacturing monitoring.
• Developing rapid, on-line analytical techniques is essential for efficient AAV production.

Purpose of the Study:

• To demonstrate the feasibility of IR-ATR spectroscopy for analyzing adeno-associated virus (AAV) type 2.
• To assess IR-ATR spectroscopy's potential for classifying full vs. empty AAV capsids.
• To explore the application of IR-ATR spectroscopy in real-time process analytical monitoring (PAT) for viral vector manufacturing.

Main Methods:

• Utilized infrared attenuated total reflection (IR-ATR) spectroscopy for AAV sample analysis.
• Employed chemometric methods, including partial least squares regression (PLS-R), for spectral data processing.
• Applied the technique to reference standard materials of full and empty AAV type 2 virus capsids.

Main Results:

• IR-ATR spectroscopy successfully classified samples containing predominantly empty capsids versus those with predominantly full capsids.
• Chemometric analysis enabled rapid and automated processing of spectral data.
• The study confirmed IR-ATR's capability to provide information on genome-containing particles and genomic titer.

Conclusions:

• IR-ATR spectroscopy, combined with multivariate data evaluation, is a viable method for analyzing AAV capsid content.
• This analytical strategy shows potential for on-line monitoring in bioreactor processes for viral vector production.
• The technique offers a rapid, cost-efficient, and non-destructive alternative to current complex AAV analysis methods.