Schematic showing in vitro assay designs for Eg5 motor studies. (A) Depiction of a fluorescence-based assay used to demonstrate purified full length Eg5 tetramers are capable of crosslinking and sliding microtubules in vitro . Unlabeled Eg5 motors bind to fluorescent, polarity-marked microtubules, causing the microtubules to slide apart. (B) Schematic showing optical trapping assay used to observe processive movement of Eg5 dimers . His-tagged motors are attached to streptavidin-coated beads through a biotinlyated PentaHis antibody. Coverslip surfaces are precoated with poly-L-lysine-graft- poly(ethylene glycol) polymers to prevent surface-induced denaturation of Eg5 at the glass interface. Polymers are biotinylated to allow the specific attachment of biotinlyated microtubules via a streptavidin linkage.