4.7. Kinase Assay

The haspin kinase assay was carried out using active recombinant haspin kinase (Millipore, Bedford, MA, USA) and the capacity of each kinase to transfer the radiolabeled phosphate from [γ-32P] ATP was measured according to the manufacturer’s instructions. In brief, active haspin kinase was co-incubated with each compound at 30 °C for 15 min. As a substrate, histone H3 (New England Biolabs) was added to each vial and then incubated at 30 °C for 15 min with [γ-32P] ATP solution in a magnesium acetate-ATP cocktail buffer (Upstate Biotechnology Inc., Lake Placid, NY, USA). The mixtures were then transferred onto p81 paper. Unreacted chemicals were washed out with 0.75% phosphoric acid, before the radiolabeled phosphate was measured using a scintillation counter.