Fig. 3
From: Optimization of chemical transfection in airway epithelial cell lines
Transgene expression and cellular viability vary with different optimization techniques tested. (A) GFP microscopy of 1HAE, 16HBE, and NCI-H292 cell lines at 48-hour post-transfection. L3000 was used as the transfection reagent for all treatments. 2X L3000 represents treating the cells with double the transfection mixture. L3000 6 h change represents replacement of the transfection mixture containing media within 6 h post-transfection. Trypsinization represents two brief rinses of 0.25% trypsin-EDTA of the cell cultures prior to transfection. Opti-MEM represents incubating cells during transfection in Opti-MEM instead of complete DMEM. Images are at 100X magnification and representative of transfections done in duplicate, with the experiment conducted twice. (B) Western blot and analysis of 1HAE, 16HBE, and NCI-H292 cultures 48-hour post-transfection against GFP (26 kDa) and β-actin (42 kDa) as a loading control. Only comparisons with p > 0.05 are shown. (C) Flow cytometry analysis for transfection efficiency represented as the percentage of cells gated positive for GFP, compared to control. Data represents two independent transfection experiments. Only comparisons with p > 0.05 are shown. (D) Alamar Blue cell viability assay at 24- and 48-hour post-transfection, normalized to either 24- and 48-hour control. (E) GFP microscopy of 1HAE, 16HBE, and NCI-H292 cell lines at 48-hour post-transfection with jetOPTIMUS. 6 h change treatment represents replacement of the transfection mixture containing media within 6 h post-transfection. (F) Western blot and analysis with GFP expression normalized to β-actin (G) Flow cytometry analysis for transfection efficiency with jetOPTIMUS. Data represents two independent transfection experiments. Only comparisons with p > 0.05 are shown. (H) Alamar Blue cell viability assay at 24- and 48-hour post-transfection with the jetOPTIMUS normalized to either 24- or 48-hour control. For all experiments, * = p < 0.05, ** = p < 0.01, *** = p < 0.001. All experiments included two technical replicates and were repeated twice