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Dye injection assay

Example results of gap junction communication competency of Cx43 constructs assayed by dye coupling analysis. GFP, Cx43FL-GFP, and Cx43dT-GFP constructs were transfected into N2A cells, a cell line which does not express Cx43 and is not dye coupled. Iontophoretic injection of sulforhodamine (red) into N2A cells transfected with GFP expressing plasmid showed no dye transfer (n=5) (A-C). In contrast, N2A cells transfected with the Cx43FL-GFP plasmid (n=17) showed abundant dye transfer between the injected cell (denoted by asterisk) and surrounding cells (D-F). However, N2A cells transfected with either the Cx43dT-GFP plasmid (n=15) (G-I) or the Cx43Y17S-GFP (n=10) (J-L) plasmid did not display any dye transfer from the site of injection (see asterisks in H, K). Quantitation of the extent of dye transfer showed only N2A cells transfected with the Cx43FL-GFP plasmid acquired significant dye-coupling (n=17) (M). Asterisks denote injected cells.
Data presented as mean +- SEM. Scale bars represent 20 um.
Figure taken from Francis RJ et al. Connexin43 modulates cell polarity and directional cell migration by regulating microtubule Dynamics (unpublished manuscript).