immuno-double-diffusion - أطلس مختبرات العرب

In immuno-double-diffusion, agar gels are poured onto slides and allowed to set; wells are then punched in the gel and the test solutions of antigen (Ag) and antibody (Ab) are added. The solutions diffuse out and where Ag and Ab meet they bind to each other, cross-link and precipitate, leaving a line of precipitation. The precipitin bands can be visualized by washing the gel to remove soluble proteins and then staining the precipitin arcs with a protein stain such as Coomassie blue. This technique may be used to determine the relationship between antigens (blue) and a particular test antibody (yellow). Three basic patterns appear. The numbers in the blue wells refer to the epitopes present on the test antigen. In reaction (1) the precipitin arcs formed between the antibody and the two test antigens fuse, indicating that the antibody is precipitating identical epitopes in each preparation (epitope 1). This does not mean that the antigens are necessarily identical; they are only identical in as far as the antibody cannot distinguish a difference. In reaction (2) the antibody preparation distinguishes the three different antigens, which form independent precipitin arcs. In reaction (3) the antigens share epitope 1 but one antigen also has epitope 2. This is the same situation as in (1), but in this case the antibody can distinguish them, by virture of being able to react against both epitopes. A line of identity forms with anti-epitope 1, with the addition of a ‘spur’ where the anti-epitope 2 has reacted with the second epitope, thus indicating partial rather than total identity between the antigen preparations.