Radioimmunoassay, enzymoimmunoassay - principles, practical use.
Laboratory measurement of specific IgE antibodies.Laboratory measurement of autoantibodies.Flow cytometry - principles, practical use.Laboratory methods of assessment of cellular immunity.Laboratory methods of assessment of humoral immunity.
Humoral analytes
• Precipitation
• Aglutination
• Turbidimetry
• Nephelometry
Imunofixation of serum
Humoral analytes
• Precipitation
• Aglutination
• Turbidimetry
• Nephelometry
TURBIDIMETRIE
NEFELOMETRIE
zdroj světla kyveta s precipitátem detektor
zdroj světla
detektor
kyveta s precipitátem
rozptyl světla
ELISA – assessment of specific antibodies
RAST
ELISA – assessment of antigen
Indirect immunofluorescence
Asessment of cellular imunity
• Number of cells - subpopulations
• Phagocytosis
• Activation of lymphocytes
Flow cytometry
Gating of lymphocytes
T and Th lymphocytes
T and B lymphocytes
Activated T lymphocytes
http://probes.invitrogen.com/resources/education/tutorials/4Intro_Flow/player.html
CD3+ T lymphocytes: mature T lymphocytes 50 - 75%
CD4+ T lymphocytes: helper T lymphocytes 30 - 60%
CD8+ T lymphocytes: cytoxic T lymphocytes 15 - 30%
CD25+ T lymphocytes: activated T lymphocytes 1 - 5%
CD19+ B lymphocytes: mature B lymphocytes 5 - 15%
CD56+ NK cells: natural killer cells 5 - 15%
CD14+ cells: monocytes
CD15+ cells: granulocytes
CD38+ cells: plasma cells
IMPORTANT LEUKOCYTE POPULATIONS
Ability to proliferateAbility to produce cytokines
TESTS OF PHAGOCYTIC FUNCTION
TESTS OF LYMPHOCYTE FUNCTION
No of granulocytesPhagocytar activity of granulocytesOxidative burst
Organ specific autoantibodies
• Anti-thyroidal AB
• Anti-pancreatic beta cells AB
• Anti-acetylcholine receptor AB
• Anti-gonadal AB
Systemic autoantibodies
• Anti-nuclear AB (ANA, ENA)
• Rheumatoid factor
• Anti-neutrophil cytoplasma AB (ANCA)
• Anti-phospholipid AB (APA)
Non-organ-specific autoantibodies in organ-localized autoimmune diseases
• ANCA (non-specific inflammatory bowel disease) //+ASCA//
• ATG (coeliac disease)
• SS-A, SS-B (Sjögren syndrome)