Technique/ Application or Precipitation Reaction
1. Ring test
2. Slide test
3. Tube test
4. Gel diffusion test
5. Immunodiffusion test
1. Ring Test
It is commonly used for detection of streptococci group.
This test is very simple for detection of antigen.
In a test tube (narrow test tube) serum is taken, over it antigen layered or poured.
A white zone is formed at the junction where the antigen -antibody reaction takes places.
For example; ASColi test in anthrax
Positive Result;
Development of white ring at the junction of antiserum and antigen solution indicates positive test.
Negative Result;
Absence of ring formation.
Serum; clear yellowish fluid obtained upon separating blood.
Antiserum; antiserum is a serum prepared from human or animal serum containing antigen specific for fight against an infectious disease.
2. Slide test
Drop of antigen and antibody are placed on slide and mixed by shaking, floccules appears indicating positive test.
For example; VDRL (veneral disease research laboratory) test for syphilis - slide flocculation.
If the test is positive; then next step to confirm the result.
1. FTA-ABS test (Fluorescent treponemal antibody absorption test); blood test
2. CSF-VDRL test to diagnose syphilis in the brain or spinal cord.
3. Tube test
Khan test for syphilis in tube flocculation test.
When the antigen solution is mixed with the solution of the antibody in a tube, floccules appear that remain suspended in a solution.
If the antigen-antibody reaction takes place, floccules appear indicating positive test.
4. Gel diffusion tests
It is usually performed in soft agar or agarose gel.
In this case the antibody incorporated in the agar gel in a test tube and antigen solution is layered over it.
The antigen diffuses downward and forms line of precipitating, where it meets the antibody.
a. Single Diffusion in one dimension
The Ab incorporated in agar gel in a test tube and Ag solution is layered over it.
The Ag diffuses downwards through agar gel forming line of precipitation. When line of precipitation is single, it is called single diffusion in one dimension.
Number of band indicates number of different antigen present.
b. Double diffusion in one dimension
Ab incorporated in gel. Above it is placed a column of agar, Ag is layered/pour over agar.
Antigen and antibody move towards each other through agar and form precipitation band where they meet.
When the precipitation band forms between two layers, it is called double diffusion.
c. Single diffusion in two dimension
(Radial immunodiffusion) (Mancini test)
Antibody is incorporated in agar gel and pour on slide.
Circular wellls are cut into the agar and antigen is introduced into the wells.
Ring shape precipitate are formed around well.
The diameter of ring is an estimate of concentration of antigen.
d. Double diffusion in two dimension
Ouchterlony test/technique
Agar gel is poured on slide or plate and wells are cut out of it.
Antibody is placed on central well and different antigens in surrounding wells.
If two adjacent antigen are identical, the line of precipitation will be formed by them.
In unrelated Ag, the lines cross react with each other.
5. Immunodiffusion test/ Gel diffusion test
a. Immunoelectrophoresis
b. Rocket electrophoresis
c. Counter immuno electrophoresis
Uses of Precipitation Reaction
1. For identification of bacteria
2. For standardization of toxins and antitoxin
3. Used for demonstration of unknown antibodies in serum. For example; VDRL
4. For identification of antigenic component of bacteria. For example; Ascoli's test
5. Used of detection of human, blood, semen etc.
(In forensic investigation)
6. Testing for blood adulteration
For example; VDRL (venereal research laboratory test).
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