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Complement Fixation test - Microbiology animations

The complement fixation test is one of the major traditional tests for the demonstration of presence of specific antigens or antibodies. It requires a veritable zoo of reagents and numerous preparatory steps. There are almost as many versions as there have been users; the microtitre version developed at the Centers for Disease Control and Prevention (LBCF Test) includes rigorous controls and is commonly used.

Fixation of Complement
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The complement fixation test (CFT) does not depend on hemagglutinating activity of the virus, but the antibodies must fix the complement, and the sera must be free of anticomplementary activity. The test has been superseded, in many instances, by newer tests, such as enzyme immunoassay (EIA).

The terminal components of the complement cascade, C789 (the membrane attack complex), can damage cell membranes in the presence of a specific antibody, which fixes complement to the cell surface. In the CFT, erythrocytes are used as the target cell, because complement-induced leakiness of the membrane can be visualized or measured calorimetrically as an increase in free hemoglobin. In the presence of specific antibodies to an infectious agent, any compliment in the system is bound, leaving no residual complement for reaction with antibodies to the erythrocytes. Thus, the presence of a specific antibody is indicated by the absence of hemolysis.

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