Motility assays are the tools of choice for the studies regarding the motility of protein molecular motors in vitro. Despite
their wide usage, some simple, but fundamental issues still need to be specifically addressed in order to achieve the best
and the most meaningful motility analyses. Several tracking methods used for the study of motility have been compared.
By running different statistical analyses, the impact of space versus time resolution was also studied. It has been found
that for a space resolution of 80 nm and 145 nm per pixel for kinesin-microtubule and actomyosin assays, respectively,
the best time resolution was ~0.9 and ~10 frame per second, respectively. A rough relationship - RatioA and RatioM - between space and time resolutions and velocity for actin filaments and microtubules, respectively, was found. The
motility parameters such as velocity, acceleration and deflection angle were statistically analysed in frequency
distribution and time domain graphs for both motors assays. One of the aims of these analyses was to study if one or two
populations were present in either assay. Particularly for actomyosin assays, electric fields varying from 0 to ~10000
Vm-1 were applied and the previous parameters and the angle between filaments motion and the electric field vector were
also statistically analysed. It was observed that this angle was reduced by ~55º with ~5900 Vm-1. The overall behaviour
of the motors was discussed bearing in mind both present and previous results and some physio-biological
characteristics. Kinesin-microtubule and actomyosin (simple and electric fields) assays were compared. Some new
experiments are suggested in order to accomplish a better understanding of these motors and optimise their role in the
applications that depend on them.