An optimizing lentiviral titer determination assay based on Raji cells

Accurate lentiviral titer determination is crucial for optimizing transduction efficiency in gene therapy. Traditional titration methods based on the human embryonic kidney 293T (HEK293T) cell line often encounter issues with fidelity and reproducibility. This study assessed the potential of suspension cell lines, such as Raji cells, as a more reliable platform for lentiviral titration.

Transduction efficiencies were compared between HEK293T cells and various suspension cell lines, including Raji, across a range of viral doses and infection conditions, both with and without infection enhancers. Lentiviral titers were quantified using quantitative polymerase chain reaction (qPCR) with primers targeting human reference genes Albumin (ALB), Poly(RC) Binding Protein 2 (PCBP2), and lentiviral sequences including Woodchuck Hepatitis Virus (WHP) Posttranscriptional Regulatory Element (WPRE) and Group-specific antigen (gag).

Raji cells demonstrated significantly higher transduction efficiency than HEK293T cells, particularly at lower viral inputs, and maintained robust infection rates at high cell densities. qPCR-based titration revealed that functional titers in Raji cells were substantially higher than those in HEK293T cells. Moreover, infection of primary T cells using Raji-derived titers showed greater sensitivity, achieving saturation at lower viral loads compared to HEK293T-derived titers.

Raji cells offer a more reliable and efficient platform for lentiviral titration compared to the conventional HEK293T-based method. This suspension cell-based approach holds potential for enhancing the scalability and consistency of lentiviral vector production in gene therapy.