Antigen-Antibody

Interaction
Precipitation test
Lab 6
Asistant Lec. Anas Yahya, Msc. Biology , Microbiology.
Asistant Lec.khetam Mohammed Msc. Biology , Immunity
5. Immunoelectrophoresis:
♦The term “immunoelectrophoresis” was first coined
by Grabar and Williams in 1953.
♦ immunoelectrophoresis is a widely used analytical
method that separates molecules based upon charge,
size and shape.
♦ Immunoelectrophoresis refers to precipitation in
agar under an electric field. It is a process of a
combination of immuno-diffusion and
electrophoresis.
♦ The main advantage of immunoelectrophoresis is
that a number of antigens can be identified in serum.
Result:
1. The presence of elliptical precipitin represents antigen-
antibody interaction.
2. The absence of the formation of precipitate suggests no
reaction.
3. Different antigens (proteins) can be identified based on the
intensity, shape, and position of the precipitation lines.

Applications:
1 - The test helps in the identification and approximate
quantization of various proteins present in the serum.
2- This method is useful to monitor antigen and antigen-
antibody purity and to identify a single antigen in a mixture
of antigens.
Rocket Immunoelectrophoresis:

●Rocket Immunoelectrophoresis is an adaptation of

radial immunodiffusion developed by Laurell.
●It is called as “rocket electrophoresis” due to the
appearance of the precipitin bands in the shape of
cone-like structures (rocket appearance) at the end of
the reaction.
●This technique is used to detect antigen-antibody
complexes.
● And to Determine the concentration of antigen in
an unknown sample.
◙ In this method antibody is incorporated in the gel at a pH
value at which the antibodies remain essentially immobile.
◙ Antigen is placed in wells cut in the gel.
◙ Electric current is then the antigen moves out of the well
and enters the agarose gel, it combines with the antibody to
form immune complex which becomes visible.
results:
●This results in formation of a precipitin line that is
conical in shape, resembling a rocket.
●A precipitation ‘rocket’ spreading out from the
loading well indicate positive reaction or specific
antigen-antibody reaction due to the presence of
antibody specific to the antigen.
●The absence of the precipitation indicates no
reaction or the absence of any corresponding
antibody – antigen.
●The height of the rocket, and its area are directly
proportional to the amount of antigen in the sample.
APPLICATION
1. Rocket electrophoresis is used mainly for quantitative
estimation of antigen in the serum.
2. Determining the concentration of a specific protein in a
protein mixture.
3. In estimation of immunoglobulin protease activity.
4. Studies dealing with antigenic relationships between
organisms.
ADVANTAGE
1. Simple, quick, and reproducible method.
2. Several unknown samples can be analyzed on a single
plate.
3. Concentrations of proteins as little as 1 µg/mL can be
measured requiring as little as 20 ng of protein to be loaded
in a well.
Thank you for
your listening