Lysates of HEK cells, transfected as indicated, were mixed with Taxol-stabilized microtubules in either 0 or 2 mM Ca++ buffer prior to sedimentation of the microtubules and microtubule-bound proteins by centrifugation (pellet, P), leaving unbound proteins in the supernatant fraction (S). Equivalent fractions of the supernatant and pellet were assayed for KHC-ECFP, milton, Miro-Myc, and tubulin.