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Introduction to Lab Ex. 23: Hemagglutination Complement Mediated Lysis Introduction to Lab Ex.23: Hemagglutination Complement Mediated Lysis Agglutination is the clumping of particulate antigens through their binding with specific antibodies. Hemagglutination is the clumping of red blood cells through their binding with specific antibodies. Thus hemagglutination is used as a observable reaction between specific antibodies with red blood cells. Complement is a series of proteins found in the serum with non-specific immune function resulting in the lysis of target cells. Properties of complement include: - non-specific immune function - inactivated by heat (560C for 30mins) - consists of 9 different proteins in a cascade - activated by: specific antigen –antibody complex formation (classical certain chemicals – endotoxin (alternate) Hemagglutination • Hemagglutination involves agglutination of RBCs. • Viral hemagglutination inhibition tests for antibodies by the antibodies' ability to prevent viruses from agglutinating RBCs. Figure 18.7 The Complement System • Serum proteins activated in a cascade. Figure 16.10 Effects of Complement Activation • Opsonization or immune adherence: enhanced phagocytosis • Membrane attack complex: cytolysis • Attract phagocytes Figure 16.11 Effects of Complement Activation Figure 16.12 Experiment: Ag=antigen; Ab= antibody; C’= complement Premise: Antigen used is sheep RBC (sRBC) Antibody used is anti-sRBC - antibody to sRBC (from immunized rabbit blood) Normal serum is from normal rabbit (Ab absent) Immune serum is from rabbit immunized with sRBC (Ab present) Normal serum (Ab absent; C’ present) Immune serum (Ab present; C’ present) Normal serum heated (Ab absent; C’ absent) Immune serum heated (Ab present; C’ absent) Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement Tube 1: sRBC + saline = (Ag) No reaction Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement Tube 2: sRBC + Normal serum = (Ag) (-Ab;+C’) No reaction Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement Tube 3: sRBC + Immune serum = (Ag) (+Ab;+C’) Hemolysis Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement Tube 4: sRBC + Normal serum (heated) = (Ag) (-Ab;-C’) No reaction Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement Tube 5: sRBC + Immune serum (heated) = (Ag) (+Ab;-C’) Hemagglutination Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement Tube 6: sRBC + Normal serum + Heated Immune serum = (Ag) (-Ab;+C’) (+Ab;-C’) Hemolysis Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement Tube 1: sRBC + saline = No reaction Tube 2: sRBC + Normal serum = No reaction Tube 3: sRBC + Immune serum = Hemolysis Tube 4: sRBC + heated normal serum = No reaction Tube 5: sRBC + heated Immune serum = Hemagglutination Tube 6: sRBC + Normal serum + heated Immune serum = hemolysis Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement Tube 1: sRBC + saline = No reaction Tube 2: sRBC + Normal serum = No reaction Tube 3: sRBC + Immune serum = Hemolysis Tube 4: sRBC + heated normal serum = No reaction Tube 5: sRBC + heated Immune serum = Hemagglutination Tube 6: sRBC + Normal serum + heated Immune serum = hemolysis