Centrifugation is a laboratory process that utilizes centrifugal force to separate or concentrate materials in liquid suspensions, particularly effective for particles smaller than 5 micrometers that don’t sediment under gravity. The document explains differential and density gradient centrifugation methods, the principles of centrifugal force, and factors influencing sedimentation including particle mass, density, and shape. Limitations of differential centrifugation are noted, particularly regarding the potential for smaller particles to pellet alongside larger ones.

1. BY: PURBASA MAITI
MSc MICROBIOLOGY

2. DEFINITION
PRINCIPLE
TYPES OF CENTRIFUGE
DIFFERENTIAL
CENTRIFUGATION
DENSITY GRADIENT
CENTRIFUGATION

3. DEFINITION
Centrifugation is a process which involves the use of the centrifugal
force for the sedimentation of heterogeneous mixtures with centrifuge
used in laboratory.
It is the process used to separate or concentrate materials suspended in
liquid medium.
The particles are suspended in liquid
medium and placed in a centrifuge
tube. The tube is then placed in a rotor and
spun at a definitive speed. Rotation of the
rotor about a central axis generates a
centrifugal force upon the particles in the
suspension.

4. PRINCIPLE
If the particle size in dispersion is 5 micro meter or less then they undergo Brownian motion
hence don't sediment under gravity. Therefore a stronger force , centrifugal force is applied
to separate the suspended particles.
Centrifugal force is a pseudo force
directed away from the axis of
rotation that appears to act on all
objects when viewed in a rotating
frame of reference..

5. If a particle spins in a centrifuge then the centrifugal force acting on it
F=centrifugal force
m= mass(kg)
v= velocity(m/s)
r= radius(m)
Same particle experience gravitational force
G= mg
Centrifugal effect (C) is the ratio of these two forces indicates how much
greater F is than G.
C= F/G

6. n= speed of rotation
d= diameter of rotation
Centrifugal effect is
directly proportional to:
Diameter of rotation
Square of Speed of
rotation
The radial force generated by the
spinning rotor is expressed relative to
the earth's gravitational force and
therefore is known as the relative
centrifugal force (RCF) or the
"g force.

7. So the sedimentation depends on:
Centrifugal force
Mass, density, volume of the particle
Density of the medium
Shape of the particle

8. Types of centrifuge

9. Differential centrifugation
Differential centrifugation is a
common procedure in microbiology
and cytology used to separate
certain organelles from whole cells .
In the process, a tissue sample is
first lysed to break the cell membranes and
mix up the cell contents.
The homogenate is then subjected to
repeated centrifugations, each time
removing the pallet and increasing the
centrifugal force.
Separation is achieved based on the size
of the particle: larger particle will sediment
first.

10. LIMITATION
The centrifugal force necessary to pellet the larger particles
from the top of the solution is also often sufficient to pallet
the smaller particles near the bottom of the tube.

11. Density gradient centrifugation
It separates the sample on the basis of
the difference in density.
Sucrose gradient is created in the
centrifugal tube
The particles of interest are placed on
the top of the tube.
On applying centrifugal force the
particle travel down the gradient until
they reach the point at which their density
matches the density of the surrounding
sucrose.