Lymphocytes
Methods to Obtain  Non-coagulating  Blood - Heparin (5-10 IU/ml): activates antithrombin III, blocks the action of thrombi...
Marker:   Cell Subpopulation CD3 , CD4, CD8   T-lymfocytes CD19, CD20, CD21  B-lymfocytes CD16/CD56  NK cells CD14/DR  mon...
CD antigens <ul><li>Antigens expressed on leukocytes  </li></ul><ul><li>Abbreviation o Custer of Differentiation </li></ul...
Flow cytometer –FACS Fluorescence-Activated Cell Scanner (Sorter)
Flow cytometer
Flow cytometry http://images.google.cz/imgres?imgurl=http://www.abcam.com/ps/CMS/Images/Flow-Cytometry-Diagram2.jpg&
lymfocytes monocytes granulocytes granularity size
CD3 CD4
CD3 CD8
CD3 CD8
Laboratory investigation of lymphocytes   <ul><li>Enumeration of lymphocyte subsets by monoclonal antibodies against typic...
Delayed-type of hypersenzitivity test
Tuberculin test
 
 
Cell-Mediated Immunity (CMI) tests <ul><li>Uses the principle of delayed type of hypersensitivity </li></ul><ul><li>Common...
Investigation of function of neutrophils <ul><li>Baktericidial killing of various bacteria of fungi </li></ul><ul><li>Chem...
NBT
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Immunology vi lab_cell_immunol

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Immunology vi lab_cell_immunol

  1. 1. Lymphocytes
  2. 2. Methods to Obtain Non-coagulating Blood - Heparin (5-10 IU/ml): activates antithrombin III, blocks the action of thrombin - EDTA: binds Ca2+ - Citrate: binds Ca2+
  3. 3. Marker: Cell Subpopulation CD3 , CD4, CD8 T-lymfocytes CD19, CD20, CD21 B-lymfocytes CD16/CD56 NK cells CD14/DR monocytes HLA-DR; CD25; CD69 activation markers The most important CD antigens of lymphocytes
  4. 4. CD antigens <ul><li>Antigens expressed on leukocytes </li></ul><ul><li>Abbreviation o Custer of Differentiation </li></ul><ul><li>Each antigen is designed : CD+ a number (CD1- CD340..) </li></ul><ul><li>The antigens were designated after they were recognized and characterized – no system exists. </li></ul>
  5. 5. Flow cytometer –FACS Fluorescence-Activated Cell Scanner (Sorter)
  6. 6. Flow cytometer
  7. 7. Flow cytometry http://images.google.cz/imgres?imgurl=http://www.abcam.com/ps/CMS/Images/Flow-Cytometry-Diagram2.jpg&
  8. 8. lymfocytes monocytes granulocytes granularity size
  9. 9. CD3 CD4
  10. 10. CD3 CD8
  11. 11. CD3 CD8
  12. 12. Laboratory investigation of lymphocytes <ul><li>Enumeration of lymphocyte subsets by monoclonal antibodies against typical cell-surface markers - CD3 (T-lymphocytes), CD4 (helper T-lymphocytes), CD8 (suppressor/cytotoxic T-lymphocytes), CD19 or CD20 (B-lymphocytes), CD16 (NK cells). </li></ul><ul><li>Evaluation of function - lymphocyte proliferation tests - determines response to various stimuli: </li></ul><ul><ul><li>specific (antigen, anti CD-3) </li></ul></ul><ul><ul><li>non-specific -polyclonal mitogens: PHA (Phytohaemagglutinin), ConA (Concanavaline A), PWM (Pokeweed mitogen) </li></ul></ul>
  13. 13. Delayed-type of hypersenzitivity test
  14. 14. Tuberculin test
  15. 17. Cell-Mediated Immunity (CMI) tests <ul><li>Uses the principle of delayed type of hypersensitivity </li></ul><ul><li>Common anamnestic antigens are used (tuberculin, candidin, trichofitin, staphylococcal antigens…) </li></ul><ul><li>In normal situation induration shout be formed by 48 hours. </li></ul><ul><li>If the patient does nor react to majority of antigens a deficiency of T- cell mediated immunity is suspected. </li></ul>
  16. 18. Investigation of function of neutrophils <ul><li>Baktericidial killing of various bacteria of fungi </li></ul><ul><li>Chemotaxis (e.g. under agarose) </li></ul><ul><li>Ingestion of inert particles </li></ul><ul><li>Metabolic activity - production of reactive metabolites of oxygen </li></ul><ul><ul><li>NBT test (reduction of colourless nitro-blue-tetrazolium into blue formasan) </li></ul></ul><ul><ul><li>Chemilu mi nescence - luminol after is oxidation spontaneously reduced and emits light </li></ul></ul>
  17. 19. NBT
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