Complement fixation test
The complement fixation test is an immunological medical test that
Purpose detects specific antigen
can be used to detect the presence of either specific antibody or
specific antigen in a patient's serum, based on whether complement or antibody
fixation occurs. It was widely used to diagnose infections, particularly
with microbes that are not easily detected by culture methods, and in rheumatic diseases. However, in clinical
diagnostics labs it has been largely superseded by other serological methods such as ELISA and by DNA-
based methods of pathogen detection, particularly PCR.
Contents
Process
Testing for antigen
Semi-quantitative testing
References
External links
Process
The complement system is a system of serum proteins that react with antigen-antibody complexes. If this
reaction occurs on a cell surface, it will result in the formation of trans-membrane pores and therefore
destruction of the cell. The basic steps of a complement fixation test are as follows:[1]
1. Serum is separated from the patient.
2. Patients naturally have different levels of complement proteins in their serum. To negate any
effects this might have on the test, the complement proteins in the patient's serum must be
destroyed and replaced by a known amount of standardized complement proteins.
1. The serum is heated in such a way that all of the complement proteins—but none of the
antibodies—within it are destroyed. (This is possible because complement proteins are
much more susceptible to destruction by heat than antibodies.)
2. A known amount of standard complement proteins are added to the serum. (These proteins
are frequently obtained from guinea pig serum.)
3. The antigen of interest is added to the serum.
4. Sheep red blood cells (sRBCs)[2] which have been pre-bound to anti-sRBC antibodies are
added to the serum. The test is considered negative if the solution turns pink at this point and
positive otherwise.
If the patient's serum contains antibodies against the antigen of interest, they will bind to the antigen in step 3
to form antigen-antibody complexes. The complement proteins will react with these complexes and be
depleted. Thus when the sRBC-antibody complexes are added in step 4, there will be no complement left in
the serum. However, if no antibodies against the antigen of interest are present, the complement will not be
depleted and it will react with the sRBC-antibody complexes added in step 4, lysing the sRBCs and spilling
their contents into the solution, thereby turning the solution pink.[1]
Testing for antigen
While detection of antibodies is the more common test format, it is equally possible to test for the presence of
antigen. In this case, the patient's serum is supplemented with specific antibody to induce formation of
complexes; addition of complement and indicator sRBC is performed as before.
Semi-quantitative testing
The test can be made quantitative by setting up a series of dilutions of patient serum and determining the
highest dilution factor that will still yield a positive CF test. This dilution factor corresponds to the titer.
References
1. "Animation Quiz 4 - Complement Fixation Test" (http://highered.mheducation.com/sites/007255
6781/student_view0/chapter31/animation_quiz_4.html). highered.mheducation.com.
2. C. Vaman Rao (2005). Immunology: a textbook (https://books.google.com/books?id=G5QZOIdq
de0C&pg=PA112). Alpha Science Int'l Ltd. pp. 112–. ISBN 978-1-84265-255-8. Retrieved
3 December 2010.
External links
Complement+Fixation+Tests (https://meshb.nlm.nih.gov/record/ui?name=Complement%20Fixa
tion%20Tests) at the US National Library of Medicine Medical Subject Headings (MeSH)
http://medmicro.atwebpages.com/cfix.htm
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This page was last edited on 26 July 2020, at 21:42 (UTC).
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