Dynamic movement of MDA -MB -231 cells in 3D culture. All MDA-MB-231 cells are GFP-labelled. (A-B) MDA-MB-231 cells cultured in 2D on a plastic dish (A) or in 3D in Geltrex matrix (B and B’, two examples). In 2D (A), almost all of the cells exhibit spindle-shaped morphology and are dissociated from each other. In contrast, the cells cultured in 3D (B, B’) are either spindle-shaped or formed aggregates. (C) Snapshots of a time-lapse movie (Additional file 1: Movie 1) of MDA-MB-231 cells cultured in 3D, showing the cells in the aggregate dissociating and migrating into the matrix. Arrows indicate cells dissociating from the aggregate. These snapshots show an enlargement of the upper area of Additional file 1: Movie 1. (D) Snapshots of a time-lapse movie (Additional file 2: Movie 2) of MDA-MB-231 cells showing elongated cells assembling and forming aggregates. Arrows indicate spindle cells becoming round and forming aggregates. These snapshots show an enlargement of the left-hand side of Additional file 2: Movie 2. (E) Snapshots of a time-lapse movie (Additional file 3: Movie 3) of MDA-MB-231 cells showing small aggregates changing location. Two aggregates are indicated by the arrow or arrowhead to follow the translocation. These snapshots show an enlargement of the centre area of Additional file 3: Movie 3. The indicated time in (C-E); hour:min. (F) 3D cultured MDA-MB-231 cells immunostained with vimentin (red) and GFP (green), showing that all cells of various cell shapes are vimentin-positive. Scale bars; A,C-F, 50 μm; B and B’, 20 μm.