1. THE COMPLEMENT SYSTEM & ITS BIOLOGICAL ROLE A Presentation By Isaac U.M. Associate Professor & HOD, Dept. of Microbiology, College of Medicine, International Medical & Technological University, Dar-Es-Salaam, Tanzania
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3. 2001 by Garland Science Figure 2.7. Schematic overview of the complement cascade. There are three pathways of complement activation: the classical pathway, which is triggered by antibody or by direct binding of complement component C1q to the pathogen surface; the MB-lectin pathway, which is triggered by mannan-binding lectin, a normal serum constituent that binds some encapsulated bacteria; and the alternative pathway, which is triggered directly on pathogen surfaces. All of these pathways generate a crucial enzymatic activity that, in turn, generates the effector molecules of complement. The three main consequences of complement activation are opsonization of pathogens, the recruitment of inflammatory cells, and direct killing of pathogens.
8. The Multiple Activities of the Complement System Serum complement proteins and membrane-bound complement receptors partake in a number of immune activities: lysis of foreign cells by antibody-dependent or antibody-independent pathways; opsonization or uptake of particulate antigens including bacteria, by phagocytosis; activation of inflammatory responses; and clearance of circulating immune complexes by cells in the liver and spleen. Soluble complement proteins are schematically indicated by a triangle and receptors by a semicircle; no attempt is made to differentiate among individual components of the complement system here.
17. Classical Pathway 2 001 by Garland Science Figure 2.14. Cleavage of C4 exposes an active thioester bond that causes the large fragment, C4b, to bind covalently to nearby molecules on the bacterial cell surface. Intact C4 consists of an α, a β, and a γ chain with a shielded thioester bond on the α chain. This is exposed when the α chain is cleaved by C1s to produce C4b. The thioester bond (marked by an arrow in the third panel) is rapidly hydrolyzed (that is, cleaved by water), inactivating C4b unless it reacts with hydroxyl or amino groups to form a covalent linkage with molecules on the pathogen surface. The homologous protein C3 has an identical reactive thioester bond that is also exposed on the C3b fragment when C3 is cleaved by C2b. The covalent attachment of C3b and C4b enables these molecules to act as opsonins and is important in confining complement activation to the pathogen surface.
22. 2 001 by Garland Science Figure 2.13. Mannan-binding lectin forms a complex with serine proteases that resembles the complement C1 complex. MBL forms clusters of two to six carbohydrate-binding heads around a central collagen-like stalk. This structure, easily discernible under the electron microscope (lower panels) has been described as looking like ‘a bunch of tulips.' Associated with this complex are two serine proteases, MBL-associated serine protease (MASP)-1 and -2. The structural disposition of MASP proteins in the complex is not yet determined. On binding of MBL to bacterial surfaces, these serine proteases become activated and can then activate the complement system by cleaving and activating C4 and C2. Photograph courtesy of K.B.M. Reid.
The test system : a sample ( patient serum , if you are searching for antibody) and specific antigen (for example, virus antigen). Step #1 – incubate serum + test antigen; if Antibodies to the test antigen are present, immune complexes will form, if not only free antigen is present Complement Step #2 Add complement: if Antibody/Antigen complexes are present, complement will bind to them and be used up = ‘fixed’; if not present then free complement remains. The indicator system : sheep red cells and hemolysin (= antibody which binds sheep red cells. Step #3 Add the indicator: anti-red blood cell antibodies equal hemolysin bind to the rbc. If free complement is available, it will bind the antibody-coated cells and lyse them Success of the assay depends on having exactly the right amount of complement present; test and indicator sera are heated to inactivate any complement present in them, if not hen the red blood cell survive. Assay can be made quantitative by measuring amount of Hb released, by spectrophotometry.
In it's simplest form the test is used to detect a patient serum antibody, so an ANTIGEN that is recognized by that antibody is the first reagent shown. If the antibody is present in the patient's serum it binds to the antigen, and the complement reagent is completely consumed in the reaction. (The test can also be used to look for antigen in the serum by modifying the reagents used). The complement fixation assay indicator system uses sheep red blood cells (SRBC) and anti-SRBC antibody. If the antibody specific for the antigen in the assay is present in the patient's serum, then complement is completely consumed in the reaction and there is none left to bind to the SRBC/anti-SRBC complexes. A Test Positive For Ab = NO HEMOLYSIS
If there is NO ANTIBODY PRESENT in the patient's serum the antigen is not bound, and the complement reagent does not have immune complexes with which to react. Complement is still present in the indicator reaction and binds strongly to the SRBC/anti-SRBC complexes. This causes the SRBCs to burst in a process called hemolysis. A Test Negative For Ab = LOTS OF HEMOLYSIS
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