Electrophoresis of individual microtubules in microchannels

Abstract

We use micrometer-sized fluidic channels to confine and measure electrophoresis of freely suspended individual microtubules. We measure orientation-dependent velocities of microtubules and the electro-osmotic flow mobility in our channels to infer the anisotropic electrophoretic mobility of microtubules under physiological conditions. We discuss the difference between electrophoresis and purely hydrodynamic motion and its implications for interpreting mobility measurements. We show that the mobility anisotropy is a factor of 0.83, clearly different from the well known anisotropy factor of 0.5 in Stokes drag coefficients for cylindrical objects. We also show that the velocity is independent of microtubule length, which would be different for hydrodynamic motion. We demonstrate that the electric force on the counterions has important consequences for the interpretation of electrophoresis experiments and that ignoring this can lead to an underestimation of the effective charge by orders of magnitude. From the electrophoresis measurements, we calculate an effective surface-charge density of −36.7 ± 0.4 mC/m² for microtubules. Electrophoretic measurements of subtilisin-digested microtubules, which have the negatively charged C termini on the outer surface removed, show a 24% decrease in mobility and, correspondingly, in surface charge, but no change in anisotropy.