Exercise 9.5 - Viscosity & Polymeriztion of Microtubules
- Tubulin (Brain extract from
- Waterbath or incubator at 37° C
- Compute the amount of GTP (M.W. 523) needed to make a final
concentration 0.5 mM GTP when added to 40 ml. of brain extract in
microtubule buffer. Likewise, compute the amount of dry ATP (M.W
507) needed to make a final concentration of 0.5 mM ATP.
- With the brain extract in an ice bath, add both the GTP and
the ATP (swirl to dissolve) and immediately transfer the mixture
to a water bath at 37° C.
- Use a viscometer to measure the viscosity at 3 minute
intervals for a period of 30 minutes, or until change is no
A viscometer is a device which makes use of capillary flow.
Insert a glass viscometer into a water bath at the appropriate
temperature (37° C for polymerization) and add a 3.0
ml sample to the viscometer. Apply suction or pressure as
appropriate to the viscometer to pull the sample into the
expanded sample chamber. Release the suction (or pressure) and
measure the time it takes for the sample to move from the upper
mark on the viscometer to the lower mark.
Use sucrose solutions of known viscosity to calibrate the
- Plot the change in viscosity with time of incubation.
Tubulin and MAP's will polymerize in the presence of Mg
and GTP. The tubulin extract is placed in a buffer that supplies
all of the necessary ingredients, except GTP. Upon the addition
of GTP, the tubulin will begin to polymerize and can be
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Cell Biology Laboratory Manual
Dr. William H. Heidcamp, Biology Department, Gustavus Adolphus College,
St. Peter, MN 56082 -- firstname.lastname@example.org