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. 2015 Dec 21;5:285. doi: 10.3389/fonc.2015.00285

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Copyright © 2015 de Groot, Hsia, Anzola, Motamedi, Yoon, Wong, Jenkins, Lee, Martinez, Davis, Gahman, Desai and Shiau.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Immunoblotting of activation loop phosphorylation of Aurora A/B and of pH3(Ser 28) and pH3(Ser 10) in HeLa cells following inhibitor treatments. (A) Analysis of MK-5108 and MK-8745. The targets blotted are indicated on the right; MW markers (in kD) are on the left. The topmost blot is with an antibody that recognizes the phosphorylated activation loops of both Aurora A and Aurora B. The protocol used to prepare mitotic taxol-arrested lysates is summarized above the blots. (B) Analysis of AZD1152-HQPA and GSK1070916 as in (A). The only difference is the protocol used to prepare the mitotic lysate; as Aurora B inhibition overrides taxol-based mitotic arrest, a synchronization procedure followed by proteasome inhibition, schematized above the blots, was used. Blots were repeated in at least two independent experiments, which were highly consistent; blot sets from one representative experiment are shown. For (B), see Figure S6 in Supplementary Material for an independent experiment performed at a higher top dose (300 nM).