96 well black untreated polystyrene microplates

Steady-state fluorescence polarization (FP) recordings were performed using a SpectraMax i3x plate reader (Molecular Devices).^{61 (link),76 (link)} All steady-state FP measurements were conducted using a buffer that contained 20 mM Tris-HCl (pH 7.5), 150 mM NaCl, 1 mM TCEP, 0.005% Tween 20, and 96-well black untreated polystyrene microplates (Corning Inc.). Other details of steady-state FP measurements were previously reported.^{44 (link)} 100 μL of each 20 nM labeled SET1_Win peptide was added to individual wells at a final concentration of 10 nM. The steady-state FP anisotropy was measured on the plates after a 1 h incubation at room temperature in the dark. WDR5-dependent dose-response data were averaged and then fitted using a four-parameter logistic function to acquire the binding affinity (K_D) for each interaction pairs.