Marked regional differences of E- and/or N-cadherin–positive AJ structures in cell cultures of hepatocellular carcinoma PLC cells. (A–D) Laser-scanning double-label immunofluorescence microscopy of reactions with antibodies to N (A, C, and D, green; B, red)- and E (A, C, and D, red; B, green)-cadherin in parts of the same PLC cell colonies, be they small (A and B) or large and near confluent (C and D). (A) Note that in the cell colony the upper portion is positive for N-cadherin only (green), whereas most of the cell–cell contact regions in the lower part of the colony are positive for both cadherins (yellow merge color). (B) Another frequent aspect of N- and E-cadherin colocalization. Close localization presents regions of merged localization (yellow) as well as regions positive for N-cadherin (red) or E-cadherin (green), the latter occurring either in small dots or whiskers or in separate, often quite extended thin zonula-like lines. (C) Dense-grown monolayer cell culture region presenting purely E-cadherin–positive (red circumscribed cells in the bottom left and the top right) or purely N-cadherin–positive (green cell–cell contacts) cells with only some plasma membrane contact regions that show E- and N-cadherin–positive (yellow) merged color cell membranes. (D) More advanced state of E–N heterodimer formation (yellow merge color) in a cell colony (green, N-cadherin; red, E-cadherin), shown at higher magnification and allowing to resolve numerous individual AJs (punctate structures). Bars, 20 µm.