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. 2019 Jan 25;13:187–196. doi: 10.1016/j.omtm.2019.01.005

Figure 1.

Figure 1

High-Density Culture of Human CD34+ Cells Enhances Transduction Efficiency with Lentiviral Vectors

(A) We transduced human CD34+ cells (1e5) with an EGFP-encoding lentiviral vector at MOI 50 after overnight pre-stimulation (100 ng/mL each of stem cell factor, fms-related tyrosine kinase 3 ligand, and thrombopoietin) among 6-well (5e4/mL), 12-well (1e5/mL), 24-well (2e5/mL), and 96-well (5e5/mL) plates with or without fibronectin coating. (B) We evaluated transduction efficiency (%GFP), GFP intensity in GFP-positive fractions, and cell counts 3 days later. (C) We then compared lentiviral transduction at various concentrations of CD34+ cells (5e5/mL, 1e6/mL, 2e6/mL, and 4e6/mL) in a 96-well plate (200 μL/well) at the same MOI 50 without fibronectin coating. We evaluated %GFP, GFP intensity, and cell counts 4 days after transduction. In addition, average vector copy number (VCN) per cell was analyzed 6 days after transduction. (D) We evaluated higher cell densities of CD34+ cell transduction (1e6/mL, 2e6/mL, and 4e6/mL) in the same vector volume as well as higher vector volumes of CD34+ cell transduction in the same cell amount (5e5/mL). Values: mean ± SE. All experiments were performed in triplicate.