Define the goal of agglutination assays

Define antibody titer

• highest dilution of Ab that causes agglutination

What causes precipitation/agglutination?

1. Formation of insoluble lattice formed between Ag and Ab at the zone of equivalence
2. forms diffusion gradients that are Ig specific, causing precipitation line

• autoimmune disease
• cancer
• infection

1. immunize animal with Ag of choice
2. make immortal B cell (hybridoma) by fusing with Ig secondary myeloma cell
3. select for specific Ag

1. incubate cells with fluorescent labeled Ab
2. run cells past laser which can excite fluorescent emission

• diagnose cancer
• CD4 T cells
  • detect Ag specific T cells with MHC/peptide complex

Process of ELISA

1. Ab coated washed
2. primary anti-Ab added
3. secondary conjugating Ab added
4. wash and add substrate
5. measure color change (intensity of color change correlates with increased Ag bound)

Clinical tests for ELISA

cytokines

Ab levels

hormone

1. bead coated with single allergen
2. add patient serum
3. incubate, wash
4. add radiolabeled anti-IgE Ab
5. measure how much anti-IgE Ab is bound

• hemolytic anemia
  • mononucleosis
  • drug induced
  • lupus associated
  • autoimmune
  • hemolytic disease of newborn

1. blood sample from patient in test tube
2. add Coomb's reagent (antihuman Ab)
3. if you have agglutination, positive for anemia

1. combine recipient with donor blood sample
2. recipient Ig's that target donor's RBC's form Ab:Ag complexes
3. add Coomb's reagents
4. if you get agglutination, positive for hemolytic disease of newborn