Complement%20

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ComplementComplement Fixation

• Hugh B. Fackrell

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Content Outline

• Complement Components
• Complement Activation
• Biological Consequences
• Complement Fixation

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Complement system
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Phases in the Complement Cascade

• Recognition
• Activation Amplification
• Membrane Attack

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Complement Cascade
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Complement Activation

• Classical pathway
• Alternative pathway
• Membrane attack complex

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Complement Components

• The Classical Pathway

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Membrane Bound Antigens
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Soluble Antigen
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C4
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C2b
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C3
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C5
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C5a C5b
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C6
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C7
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C7 becomes hydrophobic
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C8
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C8 in membrane
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C9
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C9 pore
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An Alternative Complement Pathway

• Part of Innate immunity
• Activated without Antibody

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Alternate Vs Classical Paths
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Regulation of Complement
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Biological Consequences

• Cell lysis
• Inflammation
• Opsonization
• Viral neutralization
• Solubilization of immune complexes

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Complement Fixation Test

• Fig 18.8

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Complement Fixation AssayTest Phase

• Positive serum
• antibodies
• Antigen
• Complement
• AG-AB complex formed
• Complement used up

• Negative serum
• no antibodies
• Antigen
• Complement
• No Ag-Ab complex
• Complement NOT used up

No visible difference!!!
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Complement Fixation AssayIndicator phase

• Positive Test phase
• No complement
• Indicator system
• Sheep RBCs
• Anti sheep RBCs
• Result
• NO LYSIS of RBCs
• CLOUDY

• Negative test phase
• Complement
• Indicator System
• Sheep RBCs
• Anti Sheep RBCs
• Result
• LYSIS of RBCs
• CLEAR

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CF Advantages

• Reagents easily available
• Semi quantitative or quantitative
• Wide application
• Extremely sensitive

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CF Disadvantages

• Indirect Test
• Need adequate controls
• Reagents unstable
• Quantitation time consuming
• Cannot be automated

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DONE!!!