The Complement Fixation Test

Laboratory Services Section

The following tests are done by Complement Fixation:

  • Adenovirus
  • Fungal Panel (Blastomyces, Coccicioides, & Histoplasma)
  • Influenza A & B
  • Parainfluenza 1, 2, & 3
  • Poliovirus 1, 2, & 3
  • Respiratory Syncitial Virus (RSV)



The complement fixation test is based on the use of complement, a biological substance present in the sera of normal animals. Its great value is its predictable activity in the presence of serologically reacting factors and its nonspecificity, that is, it is not like an antibody with a narrow range of reactions or increased concentration occurring in a host following immunization or infection. Furthermore, it is easily destroyed by heating at temperatures that have no deleterious effect on antibodies.

It is the nature of complement not to react with an antigen or an antibody alone but to enter into combination with antigen-antibody complexes. The lack of specificity of complement allows it to react with almost any antigen-antibody complex. Therefore, in this test procedure, unless complement is fixed by the particular antigen and antibody system in question, it will remain free to react as an indicator, resulting in a "negative"  test.  If on the other hand, complement is fixed by the antigen and antibody system under study, it is not free to react in the indicator system, resulting in a "positive" test.  The indicator system used in CF is sheep red blood cells (RBCs).  In a positive or reactive test, the complement is bound to an antigen-antibody complex, and is not free to interact with sensitized RBCs so they remain unlysed and settle to the bottom of the well to form a button.  On the other hand, in a negative or nonreactive test, the complement remains free since there is no antigen-antibody complex for it to bind to, and it interacts with the sensitized RBCs causing them to lyse.

In the Molecular and Serological Analysis Group  a specially standardized complement fixation test, the Laboratory Branch Complement Fixation Test (LBCF), is used. The LBCF is reliable only when carefully standardized throughout. All reagents involved must be used at optimal reactivity. Therefore it is imperative that all known reagents be carefully prepared and standardized to insure a completely balanced system. These standardizations, which involve titrations of sheep red blood cells, hemolysin, complement, and antigens before the test proper can be performed, render the complement fixation test rather difficult and time consuming. An actual test requires 2 days to complete.  It is this reason, as well as the fact that many of the test antigens and antisera are not available commercially, that the test is not often incorporated into the test program of the average clinical laboratory.

Complement Fixation test


Last updated September 16, 2010