APPLICATIONS OF

CENTRIFUGATION
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Cell Fractionation
Velocity sedimentation centrifugation
separates particles ranging from
coarse precipitates to sub cellular
organelles.

 In a typical cell fractionation , cells or tissues in

solution are disrupted by gentle
homogenization.
This treatment ruptures the plasma membrane
but leaves most of the organelles intact.
The homogenate is then centrifuged;
organelles
such as nuclei, mitochondria, and lysosomes
differ in size and therefore sediment at
different rates.
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 Differential centrifugation results in a rough

fractionation
of the cytoplasmic contents, which may be further
purified by isopycnic (same density) centrifugation.
In this procedure, organelles of different buoyant
densities
(the result of different ratios of lipid and protein in
each
type of organelle) are separated on a density
gradient.
By carefully removing material from each region of the
gradient and observing it with a microscope, the
biochemist can establish the sedimentation position of
each organelle
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 Particles of different density can be

separated
by isopycnic centrifugation. In isopycnic
centrifugation, a centrifuge tube is filled
with a solution, the density of which
increases
from top to bottom; a solute such as
sucrose is dissolved at different
concentrations to produce the density
gradient.
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Analytical Measurements

Dr. Nikhat Siddiqi

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Density Gradient
Centrifugation

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Zonal Centrifugation Prepared

in Density Gradient

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 After centrifugation the contents of the

tube are fractionated by drop collection
from the bottom of the tube.
In analytical centrifugation optical
techniques are used to detect the
molecules.
The fractions obtained can be assayed by
radioactivity, chemical tests, enzymatic
activity etc or a combination of these.
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Dr. Nikhat Siddiqi

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Dr. Nikhat Siddiqi

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Uses
Separation of enzymes, hormones,
RNA-DNA hybrids, ribosomal
subunits, subcellular organelles, for
the analysis of size distribution of
samples of polysomes and
lipoprotein fractions.

Dr. Nikhat Siddiqi

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Dr. Nikhat Siddiqi

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