Immunology – Complement Practical                BB2218S


The term “complement” was applied by Ehrlich to describe the activity in serum, which could “complement” the ability to specific antibody to cause lysis of bacteria (Immunology, Roitt et al, 4th ed).  

The complement system plays a crucial role in host defence against infectious agents and in the inflammatory process.  It consists of twenty plasma proteins that function either as enzymes or as binding proteins that act as a cascade, where each enzyme acts as a catalyst for the next, with C3 being the most important component.  The complement system also includes multiple distinct cell-surface receptors that exhibit specificity for the physiological fragments of complement proteins and that occur on inflammatory cells and cells of the immune system.

The consequences of complement activation are:

  • Opsonisation
  • Activation of leucocytes
  • Inflammation
  • Lysis of target cells

There are three different mechanisms that the complement system uses for recognising micro-organisms.  

The first pathway to be discovered was the classical pathway.  It is activated by the binding of antibody molecules, specifically IgM and IgG1, 2 and 3 to a foreign particle i.e. antigens.  Here, the complement proteins work together with antibodies to enhance the removal of antigen-antibody complexes from the body.

Another common pathway is the alternative pathway.  This is of major importance in host defence against bacterial infection as it is activated by invading microorganisms and does not require antibody, therefore this pathway is antibody independent.  It provides an amplification loop for the classical pathway of complement activation as one of the activated components of the classical pathway can also initiate the alternative pathway.

The third and final pathway is the lectin-mediated pathway.  This pathway is activated by the binding of a mannose-binding protein present in blood plasma to mannose-containing carbohydrates on the surface of bacteria or viruses.  


For the purpose of the experiment undertaken, I will only be considering the classical and alternative pathways from now on.


The classical pathway links to the adaptive immune system through the binding of immune complexes to C1q.  The alternative pathway, which links to the innate immune system, is activated by the chance binding of C3b to the surface microorganisms.  

Both the classical and alternative pathways lead to the formation of a convertase that cleaves C3 to produce C3a and C3b.  It is the way in which each of the two pathways lead to the production of the enzyme that cleaves the C3 that alters the pathways.  Once it has been cleaved, both pathways follow the same route.  The convertase used in the classical pathway is a combination of C 4 and C2, which forms a complex, C4b2a.  However, in the alternative pathway C3 combines with Factor B to give C3bBb.  The C3b generated by these two enzymes binds to the to the target membrane and becomes a focus for further C3b production.  This part of the cascade is called the amplification loop.  The further addition of C3b converts C4b2a and C3bBb into a C5 convertase, which catalyses the first step in the cascade that leads to the production of membrane attack complexes (MAC).

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        In the classical pathway, C1 is the first enzyme in the cascade.  It binds to the Fc region of antigen-complexed antibody molecules.   It is a pentamolecular molecule made up of one C1q, two C1r and two C1s, and is Ca2+ dependent.  Activation is initiated by the binding of C1 to complexed antibody.  The first step is the binding of antibody to two or more of the six globular domains of C1q.  This then binds to the complexed IgG or IgM causing changes in conformation of the C1 complex.  This leads to the activation of one of the C1r molecules ...

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