Objectives of Centrifugation
Separation of Components: To separate particles based on their size, shape, density, viscosity of the medium, and rotor speed.
Purification: To purify biological samples, such as proteins, nucleic acids, cells, and sub-cellular organelles.
Concentration: To concentrate particles, cells, or macromolecules from a solution.
Isolation: To isolate specific components from a complex mixture for further analysis or use.
Clarification: To remove suspended particles from a solution, thereby clarifying it.
Principle of Centrifugation
Centrifugation operates based on the principle of sedimentation, where a force greater than gravity is applied to accelerate the settling rate of particles in a sample.
When a sample is spun at high speed in a centrifuge, particles with different densities move at different rates:
Centrifugal Force:
Particles are subjected to a centrifugal force,
m is the mass of the particle,
r is the radius of rotation,
ω is the angular velocity.
Sedimentation Rate:
The rate at which particles sediment depends on their mass, the centrifugal force, and the frictional resistance they encounter from the medium they are suspended in.
Particles with higher density or larger size sediment faster towards the bottom of the tube, forming a pellet, while lighter or smaller particles remain in the supernatant.
Applications of Centrifugation
1) Biological and Clinical Laboratories:
Cell Separation: Isolating cells from blood or other body fluids.
Fractionation of Subcellular Organelles: Separating organelles like nuclei, mitochondria, lysosomes, and ribosomes.
DNA and RNA Purification: Isolating nucleic acids from cell lysates.
Protein Purification: Separating proteins based on size and density.
2) Pharmaceutical Industry:
Drug Development: Purifying drugs, vaccines, and other pharmaceutical products.
Quality Control: Ensuring the purity and concentration of pharmaceutical preparations.
3) Food and Beverage Industry:
Milk Processing: Separating cream from milk.
Wine Clarification: Removing yeast and other particulates from wine.
4) Environmental Science:
Water and Wastewater Treatment: Removing suspended solids and microorganisms from water samples.
Soil Analysis: Separating soil components for analysis of contaminants.
5) Industrial Applications:
Nanoparticle Separation: Isolating nanoparticles for use in various industrial processes.
Oil and Gas: Purifying lubricants and other fluids.
Types of Centrifugations
1) Differential Centrifugation:
Based on differences in sedimentation rates of particles.
Commonly used for separating cells, organelles, and macromolecules.
2) Density Gradient Centrifugation:
Uses a gradient medium (e.g., sucrose or cesium chloride) to separate particles based on density.
Can be further classified into isopycnic (buoyant density) and rate-zonal (size and shape) centrifugation.
3) Ultracentrifugation:
Uses extremely high speeds (over 100,000 rpm) for separating very small particles like viruses, ribosomes, and macromolecules.
Requires special equipment and is often used in research settings.