Considerations For Functional Viral Vector Titration By Flow Chemistry

By John Nguyen, DPhil, Joshua Gutierrez, Venkatesh Natarajan, PhD, and Frank Luh, Theragent, Inc.

Methods for Improving the Robustness and Reproducibility of Viral Vector Functional Titers

Accurately quantifying viral titers is a challenge in the development and manufacturing of viral vectors for cell and gene therapies. Traditional physical titers often overestimate the number of viral particles, while functional titers provide more practical information on potency and quality. However, methods like flow cytometry used to measure functional titers have their limitations.

In this white paper, we delve into the pitfalls of flow cytometry analysis and propose solutions to improve the accuracy and efficiency of functional viral vector titration. We specifically focus on lentiviral vectors (LVV) and provide insights into the considerations for functional viral vector titration using flow cytometry.

Flow cytometry's inability to distinguish between transduction by multiple viral particles and a single integration event can create challenges, leading to underestimation of the functional titer. Applying binomial and Poisson statistics to model transduction events by multiple infectious particles per cell at higher titers can help to overcome this limitation.

Additionally, we discuss other factors that can affect the accuracy of functional viral vector titration, such as false-positive gating of pseudo-transduced cells and the impact of spinfection. Adjusting gating strategies and utilizing spinfection to enhance the sensitivity and reliability of titration assays can improve accuracy adn reliability. Download the white paper to gain valuable insights into improving the robustness and reproducibility of viral vector functional titers in cell therapy manufacturing. By addressing the limitations of flow cytometry analysis and proposing solutions, you can enhance the accuracy and efficiency of viral vector titration, ultimately advancing the field of cell and gene therapies.