Complement:Measurement Secondary article Jonathan North,City Hospital NHS Trust,Birmingham,UK Artice Contents K Whaley,University of Kuwait,Safat,Kuwait The complement system comprises a group of serum proteins and cell membrane receptors that function primarily to fight infection.Clinically,measurement of Introduction complex proteolysis.Cleaved C4(C4b)will bind to the activating membr function complex. The sent an the central results of activation of these pathways are to C3 by limited pr iActivated C3(C3)will bind deposit the opsonin C3b on bacteria to promote phago cytosis,to lyse by t assembly of the terminal valently to certain surfaces (see below)and act as a high- afinity ligand for C3b receptors on phagoeytes. to the In ren :C4b2a dual com onent levels is,therefore.of value in cases of immunodeficiency inhibito and conditions that pdmheimtialswages CI-inhibitor (CI-inh).which binds to CIr and CIs and volve complemic lupus erythematosus (SIE prevents C4 activation.C4-binding protein (C4bp)dis and acts as a cofactor for the readily achieved using standard immunochemical meth ods.Nephelometry and turbidimetry can be used to assay Mannan-binding lectin activation of (CI-i h)).Enzyme-linked say (ELISA) complement or radioimmunoassay(RIA)are used to measure compo several molecules nents present in rela tively low concentrations,but s m In addition to immune c plasma centrati 0 bndheolhgemous.eobCigamdogrQhso of th jority of the uch s DNA. olecul onents To fully app eciate the condi- tions required for the functional assays,it is important to polysaccharides understand he process occurring. Accordingly In contrast,mannan-binding lectin (MBL)is a protein nditions p necess is in the dy an q when bo following section,with more detailed descriptions to be found in the individual assay sections.Full methodologies in this chapter are given in the vith two MBL-associated serine protea es(MASPs I and 2)that activate C4 and C2 to form C4b2a;the classical I here i ctly.The MR The classical pathway uire scalcium ions for its formation and binding to carbohydrate but will activate C4 in the absence of calcium once bound. ecomea that pontain imunopo on Once bound.the activated and this,in turn.activates Cls.The integrity of ENCYCLOPEDIA OF LIFE SCIENCES/2001 Nat ishing Group/www.els.netComplement: Measurement Jonathan North, City Hospital NHS Trust, Birmingham, UK K Whaley, University of Kuwait, Safat, Kuwait The complement system comprises a group of serum proteins and cell membrane receptors that function primarily to fight infection. Clinically, measurement of complement pathway activity and individual component levels is of value in cases of immunodeficiency and inflammatory conditions that involve complement activation. Introduction The complement system comprises a group of serum proteins and cell membrane receptors that function primarily to fight infection. These components interact in three activation pathways and one final pathway. The central results of activation of these pathways are to deposit the opsonin C3b on bacteria to promote phago￾cytosis, to lyse bacteria by the assembly of the terminal membrane attack complex and to promote inflammation. Clinically, measurement of pathway activity and indivi￾dual component levels is, therefore, of value in cases of immunodeficiency and inflammatory conditions that involve complement activation, the prime example of the latter being systemic lupus erythematosus (SLE). The measurement of complement protein levels is readily achieved using standard immunochemical meth￾ods. Nephelometry and turbidimetry can be used to assay levels of serum complement components that are present in relatively high concentrations (e.g. C3, C4, C1-inhibitor (C1-inh)). Enzyme-linked immunosorbent assay (ELISA) or radioimmunoassay (RIA) are used to measure compo￾nents present in relatively low concentrations, but serum/ plasma concentrations of all components have been successfully measured by ELISA. It is also possible to determine the functional activity of the majority of the complement components. To fully appreciate the condi￾tions required for the functional assays, it is important to understand the process that is occurring. Accordingly a summary of the complement pathways, stressing certain conditions necessary for activation, is given in the following section, with more detailed descriptions to be found in the individual assay sections. Full methodologies for the assays described in this chapter are given in the literature listed for further reading. The classical pathway Immune complexes that contain immunoglobulin G (IgG) or IgM can activate the classical pathway by the binding of the first component of complement (C1) via the C1q subcomponent. Once bound, the subcomponent C1r is activated and this, in turn, activates C1s. The integrity of the C1q–C1r–C1s complex is calcium-dependent. Acti￾vated C1s is a serine protease that activates C4 by limited proteolysis. Cleaved C4 (C4b) will bind to the activating complex. C2 binds to C4b when magnesium ions are present and C1s will then activate C2 to form the C4b2a complex (classical pathway C3 convertase) that activates C3 by limited proteolysis. Activated C3 (C3b) will bind covalently to certain surfaces (see below) and act as a high￾affinity ligand for C3b receptors on phagocytes. In addition to the inherent instability of C4b2a, spontaneous activation of the classical pathway is con￾trolled in the initial stages by the serine protease inhibitor C1-inhibitor (C1-inh), which binds to C1r and C1s and prevents C4 activation. C4-binding protein (C4bp) dis￾sociates the C4b2a complex and acts as a cofactor for the cleavage of C4b by factor I. Mannan-binding lectin activation of complement In addition to immune complexes, several molecules can bind the collagenous region of C1q and cause classical pathway activation. Such molecules include C-reactive protein and fibronectin, and molecules that have an anionic nature such as DNA, lipopolysaccharide and mucopolysaccharides. In contrast, mannan-binding lectin (MBL) is a protein that structurally resembles C1q itself and can activate C4 in the absence of antibody and C1q when bound to specific carbohydrates (e.g. those of many bacteria). MBL circulates as a macromolecular complex in association with two MBL-associated serine proteases (MASPs 1 and 2) that activate C4 and C2 to form C4b2a; the classical pathway C3 convertase. There is also evidence that the MBL/MASP complex can activate C3 directly. The MBL/ MASP complex requires calcium ions for its formation and binding to carbohydrate but will activate C4 in the absence of calcium once bound. Article Contents Secondary article . Introduction . Measurement of Total Complement Activity . Quantitation of Individual Components . Clinical Relevance ENCYCLOPEDIA OF LIFE SCIENCES / & 2001 Nature Publishing Group / www.els.net 1