Fig. 6. Kinetochores with weaker Bub1 staining are oriented towards the spindle pole. (A) A projection of a deconvolved image stack showing a prometaphase DLD-1 cell stained with 4B12 (anti-Bub1, red), RAA.1 (anti-aurora a, green) and Hoechst (blue). (B) An enlarged image of the region boxed in (A) showing three kinetochore pairs. The two pairs that exhibit clear Bub1 asymmetry are oriented such that the weaker staining kinetochore is closer to the pole. (C) shows a dot plot of the interkinetochore distance (K1-K2) against the fluorescence ratio (K1/K2), confirming that kinetochores exhibiting weaker Bub1 staining are generally closer to the nearest spindle pole. The scale bars represent 5 µm in (A) and 1 µm in (B).