Figure 3

Microscopic and flow cytometric analyses of double-transfected cell lines overexpressing fluorophores. (a) Simultaneous co-transfection of the sodium/bile acid cotransporter (NTCP) tagged with cyan or yellow fluorescent protein (CFP, YFP) into HEK293 cells reveals an even expression in virtually all cells. Transient co-transfection of NTCP-CFP and NTCP-YFP illustrates the heterogeneity in strength and distribution of transient transfections, compared to the stably integrated double-transfection. (b) HEK293 cells showed consistent overexpression of tdTomato and eGFP during the early selection process by live-cell imaging. (c) Flow cytometric analysis of the integration of the genes encoding eGFP and tdTomato showed a high stability over 30 passages (mean ± SEM). (d) Stability of genomic integration of simultaneously transfected CFP- and YFP-tagged sodium/bile acid cotransporter (NTCP) was additionally confirmed by microscopic analysis (mean ± SEM). Figures (c) and (d) were created using GraphPad Prism version 5.01 for Windows, GraphPad Software, La Jolla, California, USA, www.graphpad.com.