Precipitation

• • Involves combining soluble antigen with soluble antibody
• produce insoluble complex that are visible.
• Soluble antigen

  Precipitinogen
• Soluble antiboby

  Precipitin
• Visible reaction

  precipitin line
• No zone of equivalence → no precipitation reaction
• involves the interaction of antigen with antibody in correct proportion resultin in a visible precipitate

  Immunoprecipitation
• number of multivalent sites of antigen and antibody are approximately equal

  Zone of Equivalence
• Optimum concentration of antigen and antibody

  ZONE OF EQUIVALENCE
• zone of antibody excess

  PROZONE
• zone of antigen excess

  POSTZONE
• no cross-linkages are formed

  Prozone
• antigen combine with only one or two antibody molecules

  Prozone
• small agreggates are surrounded by excess antigen

  Postzone
• No lattice formation is formed

  Post zone
• cross-linkages between antigen and antibody

  Lattice formation
• Cell washing of RBCs with NSS 3x
• Scattered light that pass through the solution is Measured using.

  Nephelometry
• High Scattered light = High Antigen/antibody present
• Immunoturbidimetry


  measurement of turbidity or cloudiness of solution
• 180 degrees from an incident light

  detection device
• Measures the reduction of light intensity caused by reflection, absorption, or scatter of light as immune complex are formed

  Immunoturbidimetry
• measures scattered light at a particular angle from the incident beam as it passes through a suspension

  Nephelometry
• 30-90 degree from the light source

  Nephelometry
• amount of light scatter is PROPORTIONAL to size, shape, and concentration of molecules in the solution
• in Rate nephelometry, the px. serum contains antigen
• in rate nephelometry, the reagent contains the antibody
• Rate nephelometry used to measures:

  • IgG
  • IgA
  • IgM IgE • Kappa and lambda light chains • Complement components o C3 and C4 o Ceruplasmin o CRP o Haptoglobin
• Rate nephelometry used to measures:

  • IgG
  • IgA
  • IgM
  • IgE
  • Kappa and lambda light chains
  • Complement components
  • C3 and C4
  • Ceruplasmin
  • CRP
  • Haptoglobin
• purified high-molecular weight complex polysaccharide derived from seaweed

  Agarose
• single diffusion technique

  Migration of antigen only

  Radial Immunodiffusion
• antibody is in the support gel, and antigen is applied in a well cut in the gel
• antigen diffuses out and combines with specific antibody in the agarose to forms precipitate rings
• 2 techniques of RID

  • Mancini or endpoint method
  • Fahey or kinetic method
• antigen is allowed to diffuse to completion

  Mancini or Endpoint
• The wider the diameter, the more Ag concentration
• X axis = antigen standard


  Mancini or End point
• Y-axis = ring diameter

  Mancini or Endpoint
• • ring diameter readings taken 19 hours before equivalence.

  Fahey or Kinetic Method
• X axis = diameter


  Fahey or Kinetic