-L9 cells have a normal karyotype and form acini in three dimensions. (A) 184-hTERT-L9 cells were arrested in metaphase using colcemid prior to multicolour fluorescence in situ hybridization analysis. Metaphase spreads had a normal diploid chromosome complement devoid of numerical or structural changes (n = 7). (B) Cells cultured in three-dimensional Matrigel were fixed after 21 days and probed with antibodies targeting GM130 (apical polarity), CD49f (basal polarity) and MUC1 (luminal marker). Either Oregon Green–labelled phalloidin or an antibody targeting E-cadherin was used as a counterstain, with DRAQ5 nuclear stain. For viability assessment, unfixed structures were incubated for 20 minutes with calcein AM, ethidium homodimer 1 (EthD-1) and Hoechst 33452 immediately prior to being imaged on a Nikon confocal laser scanning microscope.