1. Introduction to Lab Ex. 23: Hemagglutination
Complement Mediated Lysis

2. 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
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.

3. 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)

5. Hemagglutination involves agglutination of RBCs.
Viral hemagglutination inhibition tests for antibodies by the antibodies' ability to prevent viruses from agglutinating RBCs.
Figure 18.7

6. Serum proteins activated in a cascade.
The Complement System
Serum proteins activated in a cascade.
Figure 16.10

7. Effects of Complement Activation
Opsonization or immune adherence: enhanced phagocytosis
Membrane attack complex: cytolysis
Attract phagocytes
Figure 16.11

8. Effects of Complement Activation
Figure 16.12

9. 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)

10. Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement
Tube 1: sRBC + saline =
(Ag)
No reaction

11. Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement
Tube 2: sRBC + Normal serum =
(Ag) (-Ab;+C’)
No reaction

12. Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement
Tube 3: sRBC + Immune serum =
(Ag) (+Ab;+C’)
Hemolysis

13. Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement
Tube 4: sRBC + Normal serum (heated) =
(Ag) (-Ab;-C’)
No reaction

14. Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement
Tube 5: sRBC + Immune serum (heated) =
(Ag) (+Ab;-C’)
Hemagglutination

15. Legend: Ag = sRBC; Ab = antibody to sRBC; C’ = complement
Tube 6: sRBC + Normal serum + Heated Immune serum =
(Ag) (-Ab;+C’) (+Ab;-C’)
Hemolysis

16. 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

17. 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