FCM analysis of transgene expression following EGFP mRNA transfection in different types of DCs.

(A) Immature Mo-DCs were cultured with GM-CSF and IL-4 and transfected at day 6 of culture with control (Melan-A) or EGFP mRNA by lipofection (bottom) or electroporation (top) and analyzed by FCM one day after transfection. The dot plots show EGFP fluorescence on the x-axis and ethidium bromide staining on the y-axis. Gates were drawn based on control mRNA–lipofected or electroporated Mo-DCs. Percentages of dead cells (upper left corner) and viable EGFP⁺ cells (lower right corner) are indicated. Results are representative of 8 independent experiments. (B) Monitoring of EGFP mRNA expression and cell viability in Mo-DCs following mRNA electroporation in function of time (n = 2). (C) 34-DCs (bottom) and 34-LCs (top) were cultured as described in “Materials and methods” and transfected at day 12 and day 25 of culture, respectively, with control (Melan-A) or EGFP mRNA by mRNA electroporation. FCM analysis was performed 24 hours after mRNA electroporation. The dot plots show EGFP fluorescence on the x-axis and ethidium bromide staining on the y-axis. Gates were drawn based on control mRNA-electroporated Mo-DCs (left). Percentages of dead cells (upper left corner) and viable EGFP⁺ cells (lower right corner) are indicated. Results are representative of 4 independent experiments.