Centrifugation

Centrifugation is a process that involves the use of the centrifugal force for the
separation of mixtures, used in industry and in laboratory settings. In chemistry and biology,
centrifugation increases the effective gravitational force on a mixture in a test tube, to rapidly
and completely bring the precipitate ("pellet") to the bottom of the tube. The
remaining solution is called the "supernate," "supernatant," or supernatant liquid. The
supernatant liquid is then separated from the precipitate by decantation or withdrawal with a
Pasteur pipette.
The equipment used for centrifugation is called a centrifuge, and the vessel that spins the
samples is called a rotor. Generally, a motor causes the rotor to spin around a fixed axis,
applying a force perpendicular to the axis. The centrifuge works using the sedimentation
principle, where the centripetal acceleration is used to separate substances of greater and lesser
density.
There are many different kinds of centrifuges, including those for very specialized
purposes. In the chemical and food industries, special centrifuges can process a continuous
stream of particle-laden liquid.
English military engineer Laval (1707-1751) invented a whirling arm apparatus to
determine drag, and Antonin Prandl invented the first centrifuge in order to separate cream from
milk to make it easier to churn butter.
History
By 1923 Theodor Svedberg and his student H. Rinde had successfully analyzed largegrained sols in terms of their gravitational sedimentation. Sols consist of a substance evenly
distributed in another substance, also known as a colloid. However, smaller grained sols, such as
those containing gold, could not be analyzed.To investigate this problem Svedberg developed an
analytical centrifuge, equipped with a photographic absorption system, which would exert a much
greater centrifugal effect. In addition, he developed the theory necessary to measure molecular
weight. During this time, Svedbergs attention shifted from gold to proteins.

Differential Centrifugation
If you had sufficient time and a vibration-free environment, you could patiently wait and
the force of gravity would bring most suspended particles to the bottom of a centrifuge tube. The
smallest particles would probably stay in suspension due to brownian motion, and most
macromolecules would be uniformly distributed because they would be in solution rather than
suspension. I don't know about you, but I don't have the kind of patience needed in order to rely
solely on gravity for separation of solid from liquid components. Besides, for practical purposes
the pellet you obtained would be way too easily disrupted for effective separation of solid

material from supernatant. Gravity would not be a terribly effective way of separating suspended
materials based on size or other characteristics.
Density gradient centrifugation using tubes is the most widely employed technique for
separating cells and cell organelles and for isolating cellular macromolecules. However, although
it is one of the cell biologists most valuable tools, it is not without disadvantages, as the amount
of material that can be fractionated in a single tube is so small.
When large quantities of sample must be fractionated (to isolate sparse organelles such as
lysosomes or peroxisomes), a very large number of tubes and gradients is needed. Much larger
quantities of sample may be fractionated using zonal rotors.
A zonal rotor consists of a large cylindrical chamber subdivided into a number of sectorshaped compartments by vertical septa (or vanes) that radiate from the axial core to the rotor
wall. The entire chamber is used during centrifugation and is loaded with a single density
gradient, each sector-shaped compartment serving as a large centrifuge tube.
The large chamber capacity of these rotors (typically 1 and 2 liters) eliminates the need
for multiple runs and multiple density gradients.
Centrifuges can be divided into types based on their rotor design: fixed angle,
swinging bucket and continuous flow. Choice of centrifuge rotor and lid will depend on
application. Fixed-angle centrifuges hold the sample containers at a constant angle
relative to the central axis and are used primarily for differential centrifugation. Swinging
head (or swinging bucket) centrifuges, have a hinge where the sample containers are
attached to the central rotor and are used primarily for gradient work. Continuous flow
centrifuges don't have individual sample vessels and are used for large volume batch
separations. Look for rotors that are easily attached, exchangeable and allow access for
cleaning. Be sure to check if your intended rotor is compatible with your centrifuges
manufacturer.
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Types of Rotor Centrifuges

Swing-Bucket Rotors

A swing-bucket rotor usually supports samples ranging in volume from 36 mL

to 2.2 mL. Swing-buckets can support two types of separations: rate-zonal and

isopycnic. Swing-buckets are preferred for rate-zonal separations, because the

distance between the outside of the meniscus and the outside of the bottom of
the tube is long enough for separation to occur.

Fixed-Angle Rotors

Fixed-angle rotors are usually used for pelleting applications to either pellet
particles from a suspension and remove the excess debris, or to collect the
pellet. Rotor cavities range from 0.2 mL to 1 mL. The most important aspect in
deciding to use a fixed-angle rotor is the K factor. The K factor indicates how
efficient the rotor can pellet at maximum speed. The lower the K factor, the
higher the pelleting efficiency.