Xeno-free production of lentiviral particles: implementing hPL as FBS replacement in cell culture.

Commonly used for delivering fluorescent or bioluminescent reporters to target genes, lentiviral vectors are an "easy-to-clone" vehicle to study up- and downregulation of gene expression induced by drug inhibitors, cytokine stimulants, chemicals, and environmental toxicants. To produce viral particles safely, several plasmid constructs are delivered to human embryonic kidney cells (HEK293-T), which use the cellular machinery to package and assemble non-replicative lentiviruses with high efficiency. However, the cell culture of HEK293-T cells still largely relies on the use of fetal bovine serum for cell survival and proliferation. At large scale, this cell culture practice consumes a great amount of animal-derived serum, driving the need for continuous farming of animals whose fetuses would be used to obtain serum, posing grave ethical and practical concerns for human-relevant research. In this study, we present a feasible and scalable method to adapt the culture medium formulation for HEK293-T cells and demonstrate how virus production responds to this cell culture adaptation. Using a commercially available platelet-based supplement derived from human blood byproducts, we maintain cell viability and proliferation rates, with up to 82% confluence, and can produce lentiviruses (approximately 10