Centrifugation- Dr M M Mir

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CENTRIFUGATION:

CENTRIFUGATION بسم الله الرحمن الرحيم Ref. Book : TEITZ text book of Clinical Chemistry and Molecular Diagnostics 4 th Edition

Definition:

Definition Centrifugation : It is the process of using centrifugal force to separate the lighter portions of a solution, mixture or suspension from the heavier portions . The equipment which is used for this process is called a centrifuge. Centrifugal Force: The outward force on a body moving in a curved path around another body[ from Latin centrum "center" and fugere "to flee")

Applications :

The applications of centrifugation in a clinical laboratory are : It helps in the removal of cellular elements from blood to provide cell free plasma or serum for analysis. It helps to concentrate cellular elements and other components of biological fluids for microscopic examination or chemical analysis. Applications

Applications :

It is used for removal of chemically precipitated proteins from specimens. It helps in the separation of protein bound or antibody bound ligands from free ligands in immunochemical and other assays. It hepls in separation of lipid components like HDL, LDL etc. Applications

Principle of Centrifugation:

Principle of Centrifugation The force required to separate the two phases by a centrifuge is called as RCF [ relative centrifugal force or relative centrifugal field]. The units are expressed as number of times greater than gravity. e.g 500 x g RCF is calculated as RCF = 1.118 x 10 -5 x r x rpm Where 1.118 x 10 -5 is a factor r is radius in centimeters from the centre of rotation to the bottom of tube during centrifugation rpm is the speed of rotation of rotor in revolutions per minute

Components of Centrifuge:

Components of Centrifuge The major components are of centrifuge are: Motor, Drive Shaft and Head or Rotor The components which control the centrifuge functions are: Power switch, Timer, Speed control, Tachometer and Brake Optional Refrigeration unit Alarm unit Safety latch

Types of Centrifuges:

Types of Centrifuges Four types of centrifuges: Horizontal head or swinging bucket Centrifuge Fixed angle or angle head Centrifuge Ultracentrifuge Axial Centrifuge

Types of Centrifuges:

Types of Centrifuges Horizontal head or swinging bucket Centrifuge: In this type, the tubes assume a horizontal plane when the rotor is in motion and a vertical position when it is at rest. During centrifugation particles travel in a constant manner along the tube while the tube is at right angles to the shaft of centrifuge. Thus the sediment is distributed uniformly against the bottom of the tube. The surface of the sediment is flat and remains so with a column of liquid on it, when the tube stops.

Centrifuge rotors:

Centrifuge rotors Fixed-angle axis of rotation At rest Swinging-bucket g Spinning g

Types of Centrifuges:

Types of Centrifuges Fixed-Angle or Angle-Head Centrifuge: In this type, the tubes are held at a fixed position at angles from 25 to 40 O to the vertical axis of rotation . Upon centrifugation, the particle are driven outward horizontally but strike the wall of the tube so that the sediment packs against the side and bottom of the tube with surface of the sediment parallel to the shaft of the centrifuge. As the rotor slows down and stops, the gravity causes the sediment to slide down and usually a poorly packed sediment is formed.

Types of Centrifuges:

Types of Centrifuges Ultracentrifuge[UC] An ultracentrifuge is a very high speed centrifuge that usually uses fixed head rotors . The most common application of an ultracentrifuge in he clinical laboratory is the separation of lipoproteins The ultracentrifuge are refrigerated as a lot of heat is generated during long operational periods. Ultracentrifuges are available in both analytical and preparative models. Speeds in the range of 100,000 to 300,000g are achieved in UC.

Types of Centrifuges:

Types of Centrifuges Axial Centrifuge An Axial Centrifuge is based on the concept that allows tubes of blood to be spun in a vertical orientation as opposed to the horizontal orientation used in traditional centrifuges.

Special Centrifuges:

Special Centrifuges Airfuge: A special class of centrifuges in which the rotor is suspended and driven by a stream of air. These can reach over 100,000 rpm in very short times but are limited to very small sample sizes. Microfuge: Another form of centrifuge, very common in laboratories now, is the microcentrifuge or microfuge . These are simple machines used with 0.5 or 1.5 ml disposable plastic tubes. Most of these machines are single speed and generate between 10,000 and 13,000 rpm.

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