Centrifugation

Dr Engelbert Buxbaum engelbert_buxbaum at hotmail.com
Mon Dec 13 07:15:37 EST 2004


ugur wrote:

> Cell extracttion procedures for protein assays have quite wide range of
> centrifugation conditions. For example one of them is at 14 000 RPM and
> the other is at 100 000 g. What are in the supernatant and in pellet in
> each case? Why there is variation? (both of the procedures for
> telomerase extrcation. 

Cells contain several organelles, which have different sedimentation
rates. Hence crude preparations of organelles can be prepared by
spinning the homogenisate at different velocities for different times:
unbroken tissue 10 min 500 g, nuclei + plasma membranes 10 min 3000 g,
mitos, plastides and heavy microsomes 30 min 20000 g and light
microsomes 1 h 100000 g. Anything that is in the supernatant after the
last spin is called "cytosol". 

In this context "g" means the centrifugal force as a multiple of the
gravitational force of the earth (g = 9.81 m s^{-2}). The centrifugal
force is calculated from the angular velocity of the rotor and its
radius. Thus in the scientific literature you will find g rather than
rpm values, because g values are comparable between different rotors.
For even more precise comparisons the pelleting efficiency of the rotor
(expressed as k-value) needs to be taken into consideration, to account
for the different pelleting distances in different rotors.



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