Fig. 2. Orientation of MTs as MCF-7 cultures reassemble cell-cell contacts. MCF-7 cells were fixed at confluence (untreated), immediately after incubation with 4 mM EGTA (-Calcium), or 15-90 minutes after replenishment of 5 mM extracellular Ca²⁺. One set of cells were treated with nocodazole (100 nM, added 60 minutes prior to experiments and maintained in media for the duration of the experiments), whereas control cultures were grown in standard media supplemented with carrier (DMSO) alone. E-cadherin and ß-tubulin were identified by dual-label epi-illumination immunofluorescence microscopy. The right-hand panel of each pair represent high-power details of the regions marked by the boxes. The density of MTs at contacts was quantitated by counting the number of MTs found within 3 µm of the E-cadherin-stained contacts. Data are means ± s.e.m., n=40, in control (C) or nocodazole (N)-treated cells. Prominent (or pioneer) MTs appeared to terminate in these cadherin accumulations (arrow), which became more numerous as cell contacts extended. Treatment of MCF-7 cells with low-dose nocodazole (100 nM) inhibited the formation of these pioneer MTs at reforming contacts.