Question

What biological techniques use the properties of antigen-antibody binding? Describe two in detail.

Answer 1

Antigen - antibody binding is a specific chemical interaction between antibodies produced by the white blood cells and antigens during immune reactions. Antigen - antibody interaction is used in precipitation reaction (soluble antigens combine with soluble antibodies in presence of an electrolyte at suitable temperature and pH to form insoluble visible complex.) and agglutination reaction ( It acts on antigen - antibody reaction in which the antibodies cross-link particulate antigens resulting in the visible clumping of the particle).

Two of the biological techniques in detail are as follows:-

1. Radial immunodiffusion (RID)

OR Mancini method is a single diffusion technique whereby a solution containing the antigen is placed into the wells in a gel or agar surface evenly impregnated with antibody. The diameter of the ring that precipitates around as a result of antigens antibody reaction corresponds to the amount of antigen in the solution.

Objectives: To determine antigen antibody complexes and the relative concentration of antigens.

Principle: Radial immuno-diffusion is a type of precipitation reaction. It is thus based on the principles of the precipitin curve which states that antigen-antibody interact forming visible cross-linked precipitate when the proper ratio of antigen to antibody is present. In the test, antibody is incorporated into agar and poured into a glass plate to form a uniform layer. Circular wells are cut into the agar and antigen is introduced into the wells. Specific antigens to the impregnated antibodies diffuse through the agar in all directions from the well and react with the antibody present forming visible precipitate or a precipitin ring. Ring shaped bands of precipitates from concentrically around the well indicating reaction. The diameter of the precipitate ring formed, corresponds to the amount of antigen in the solution.

Procedure:

1. An agar containing an appropriate antiserum (antibody) is poured in plates.
2. Carefully circular wells are cut and removed from the plates.
3. A single or series of standards containing known concentration of antigen are placed in separate wells, while control and “unknown” samples are placed in other remaining wells.
4. As the antigen diffuses radially, a ring of precipitate will form in the area of optimal antigen – antibody concentration.
5. The ring diameters are measured and noted.
6. A standard curve is prepared using the ring diameters of the standards versus their concentrations. This curve is then used to determine the concentration of the control and unknown samples. 2. ​​​​​​​ABO Blood group test:   Principle: ​​​​​​​Blood groups refer to the presence on human red blood cells of certain antigens, the blood group factors. One very important group of factors present on the red blood cells is the ABO system. The ABO group of a person depends on whether his/her red blood cells contain one, both, or neither of the 2 blood group antigens A and B. There are, therefore, 4 main ABO groups: A, B, AB and O. Antibodies (agglutinins) for the antigens A and B exist in the plasma and these are termed anti-A and anti-B. The corresponding antigen and antibody are never found in the same individual since, when mixed, they form antigen-antibody complexes, effectively agglutinating the blood.      Procedure: One end of a slide is labelled Anti-A, and the other Anti-B. A drop of Anti-A test serum is added to the end marked Anti-A, and a drop of Anti-B serum is added to the end marked Anti-B.   One drop of blood is added to each end of the slide, and mixed well, using separate wooden sticks. The results are read directly from the slide. The subject is blood group A if agglutination occurred with the Anti-A test serum; group B if agglutination occurred with the Anti-B test serum; group AB if agglutination occurred with both test serums, and O if there was no agglutination in either case. In the sample to the right, we conclude the subject has type A blood.