Counterimmunoelectrophoresis (CIE): Principle, Procedure, Uses

Counterimmunoelectrophoresis (CIE) is a modification of the Ouchterlony method that speeds up the migration of an antigen and antibody by applying an electrical current.

Principle of CIE Test

Most bacterial antigens are negatively charged in a slightly alkaline environment, whereas antibodies are neutral. When an electric field is applied in an electrophoresis apparatus filled with buffer (pH 8.4) and containing known antibodies and unknown antigens in the agarose well, the antibodies will migrate towards the negative end. In contrast, antigens will migrate towards the positive end. 

As the antibody and antigen move toward each other in an electric field, they will soon meet in optimal proportion (a zone of equivalence is formed) at some points between the well, and visible precipitation occurs.

Salient features

1. Modification of the Ouchterlony method.
2. An unknown bacterial antigen and a known specific antibody move towards each other in an electrical field.
3. Most bacterial antigens are negatively charged  (exceptions Streptococcus pneumoniae serotypes 7 and 14) in a slightly alkaline environment.
4. When Ag and Ab meet in optimal proportions, a line of precipitation appears within the agar matrix.
5. The entire procedure takes about one hour.
6. CIE can test any antigen for which antisera is available
7. Sensitivity less than that of particle agglutination (0.01-0.05 mg/ml) of antigen.

Uses of Counter-immunoelectrophoresis

Counterimmunoelectrophoresis is used to detect the presence of bacterial (H. influenzae, N. meningitidis, S. pneumoniae) or fungal (Cryptococcus neoformans) capsular polysaccharide antigens in cerebrospinal fluid (CSF).

Limitations of CIE Test

1. CIE is more expensive than agglutination-based tests.
   • Initial capital outlay for the apparatus
   • Need for large quantity of Ag and Ab