Fig. 9. Slow aggregation, compaction and decompaction assays on various related HCT-8 colon cancer cells. (A) Representative immunofluorescent staining for -catenins in the cell lines used: no expression in untransfected HCT-8/R1 cells (R1), strong expression of E-catenin in untransfected HCT-8/E8 cells (E8), and moderate expression of exogenous T-catenin in transfectant HCT-8/R1/T31 (T31). (B) After slow aggregation of single-cell suspensions for 24 hours on semi-solid agar, images of representative cultures were taken. No aggregation is seen in cultures of HCT-8/R1 cells. Cells expressing exogenous T-catenin form compacted large aggregates, similar to those of cells expressing endogenous E-catenin. (C-D) Compaction-decompaction assays. (C) After Gyrotory shaking of suspended cell cultures for 3 days, cell aggregates were similar to the results obtained in (B). (D) Small spheroids of HCT-8/R1 cells were dissociated by repeated pipetting, whereas no decompaction could be seen of larger spheroids of HCT-8/E8 and HCT-8/R1/T31 cells. (E) Quantification of compaction/decompaction by volume distribution in function of the particle diameter: unbroken line, before compaction; dashed line, compaction; dotted line, decompaction after pipetting.