Biology Reference
In-Depth Information
FIGURE 2.3
Gliding assay calculation of microtubule trajectories and persistence length. (A) TIRF image of
microtubules sparsely decorated with fluorescent GTP analogs, taken following
Section 2.2
.
Scale bar is 5 mm. (B) Microtubule trajectories from (A) reconstructed as described in
Section 2.3
. Where these trajectories appear to cross, timing information is associated with
each trajectory, permitting unique trajectory identification. (C) Average of cos
y
s
as a function
of path length,
, (dots) and fit to Eq.
(2.1)
(solid line). The data shown are an average across
all trajectories in (B). Fewer independent data for long trajectory lengths lead to statistical
fluctuations. The persistence length for these microtubules is 150 mm, consistent with very
short tip-length microtubules.
s
After individual fluorophore trajectories are calculated, the trajectories of all
fluorophores on one microtubule are automatically combined into a single microtu-
bule trajectory, with ordered positions and times. The software we have developed
for this task simply takes one fluorophore trajectory and, for each point in that tra-
jectory, searches for all other trajectories which come within a micron of that point—
any trajectories meeting this criteria are candidate trajectories. Crossing trajectories,
