Size of
pDNA/PEI particles dictates their transfection efficiency
in LVV production cells. (a) Schematic of preparation of pDNA/PEI
particles and transfection process for production of LVVs. Each exclamation
mark indicates a critical process control parameter influencing transfection
outcomes. (b) Transfection efficiencies, characterized as transgene
expression levels of the luciferase reporter, in a monolayer culture
of HEK293T cells as a function of pDNA concentration at the mixing
step and incubation time (10 s to 60 min) before dosage. For the group
of mixing at a DNA concentration of 5, 10, or 20 μg/mL, the
particles were diluted 5, 10, or 20 times, respectively, to 1 μg/mL
to dose cells. (c) Growth in the average size (z-average
diameter given by dynamic light scattering, DLS) of pDNA/PEI particles
following mixing of pDNA and PEI solutions in Opti-MEM. The growth
kinetics is dependent on the concentration of pDNA. The error bars
were derived from three independent experiments, demonstrating reproducibility
and predictability under the experimental conditions used. (d) Direct
correlation between transfection efficiency and the z-average particle size based on data points from all experiments
from (b) and (c) with varying pDNA concentrations and incubation times.