Fig. 5

The anti-CD98hc-DM1 antibody provokes cell cycle arrest in mitosis and mitotic catastrophe. A Effect of anti-CD98-DM1 (10 nM, 24 hours) on the morphology of MDA-MB231 and HS578T cells grown as monolayers. The images were taken at 10X magnification. B Quantitative analyses of the action of anti-CD98hc-DM1 on the distribution of the different cell cycle phases in MDA-MB231 and HS578T cell lines. C MDA-MB231 and HS578T cells were treated with anti-CD98hc-DM1 (10 nM) and lysed at the indicated times. Analyses of the amounts of the different proteins studied were performed by Western blotting. GAPDH was used as loading control. D Effect of anti-CD98hc-DM1 on spindle assembly and organization. MDA-MB231 cells seeded on coverslips were treated with CD98hc-DM1 (10 nM) for 24 hours, fixed and stained. Scale bars are indicated. E Detection of giant multinucleated cells after anti-CD98hc-DM1 treatment. MDA-MB231 cells were treated with 10 nM anti-CD98hc-DM1 for 48 hours, fixed and stained for nucleoporin p62 (red) and DNA (blue). Scale bar = 7.5 μm. F Quantitation of abnormal mitoses, normal mitoses and interphase cells from the experiment shown in (D). G Bar graph representation of the percentage of viable (Annexin V-negative/PI-negative) and non-viable MDA-MB231 cells at 48 and 72 hours of treatment with 10 nM anti-CD98hc-DM1. H Effect of anti-CD98hc-DM1 on the levels of several apoptosis-related proteins. MDA-MB231 and HS578T cells were treated with 10 nM anti-CD98hc-DM1, lysed at 0, 1, 2, or 3 days and the indicated proteins analyzed by Western