Fig. 4. Punctate structures are resistant to Triton X-100 extraction. Confocal immunofluorescence microscopy was performed on live C6R cell monolayers expressing wild-type and mutant Cx43-eYFP constructs, as indicated after incubation in HBSS (A,C,E,G,I,K,M) and after incubation for 15 minutes in HBSS containing 1% Triton X-100 (B,D,F,H,J,L,N) on the same field. Significant numbers of puncta remained after Triton X-100 extraction for the wild type and the various mutants. Although F52dup did not form plaque structures, some of the cell surface signal was resistant to extraction. Real-time monitoring of Triton X-100 extraction revealed that signal extracted was nearly instantaneously after addition of Triton. Overlays of images cannot be created because of an apparent mild deformation of cell architecture by Triton X-100 in non-fixed cells. Scale bar, 10 µm.