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AUTOANTIBODIES IN THE DIAGNOSIS OF AUTOIMMUNE IMMUNOPATHOLOGICAL DISEASES

AUTOANTIBODIES IN THE DIAGNOSIS OF AUTOIMMUNE IMMUNOPATHOLOGICAL DISEASES. AUTOIMMUNE IMMUNOPATHOLOGICAL DISEASES. * result of loss of autotolerance * inflammation * organ specific * organ non nespecific (systemic). AUTOANTIBODIES:.

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AUTOANTIBODIES IN THE DIAGNOSIS OF AUTOIMMUNE IMMUNOPATHOLOGICAL DISEASES

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  1. AUTOANTIBODIES IN THE DIAGNOSIS OF AUTOIMMUNE IMMUNOPATHOLOGICAL DISEASES

  2. AUTOIMMUNE IMMUNOPATHOLOGICAL DISEASES * result of loss of autotolerance * inflammation * organ specific * organ non nespecific (systemic)

  3. AUTOANTIBODIES: * the presence of autoantibodies is usually associated with immunopathology * intensity of damage is usually associated with the amount of autoantibodies * autoantibodies are usuallydirected to immunodominant epitopes

  4. CAVE: * the are healthy persons with the presence of autoantibodies * the are personswithmanifested immunopathological disease, in whom expected autoantibodies are undetectable * low level of autoantibodies (IgM) could be associated with infection *crossreactivities

  5. LABORATORY DETERMINATION OF AUTOANTIBODIES: Based on immunochemical detection by: * agglutination * precipitation: * ELISA * turbidimetry * immunobloting * immunofluorescence determination of autoantibodies is: * qualitative * quantitative: * titr * arbitrary units * concentration (standard)

  6. MOLECULAR TARGETS FOR RHEUMATOID FACTORS V domains -S-S- -S-S- -S-S- -S-S- -S-S- -S-S- -S-S- -S-S- -S-S- -S-S- -S-S- C1 -S-S- binding sites for RF Agal IgG C2 Asn 297 C domains -S-S- -S-S- C3 His 435 -S-S- -S-S- GA – specific RF

  7. IMMUNOPATHOGENESIS OF RHEUMATOID ARTHRITIS binding of RF to immunocomplex deposition of immunocomplexes to joint synovia complement RF IgG bound to collagen collagen plasma cell

  8. IMMUNOFLUORESCENCE DETERMINATION OF AUTOANTIBODIES: detection of autoantibodies reacting with tissue (intracellular) antigens Using : * tissue cuts (organ-specific autoantibodies) * cell suspensions: * Hep-2 cell line (ANF) * granulocytes (ANCA)

  9. IMMUNOFLUORESCENCE DETERMINATION OF ANTINUCLEAR AUTOANTIBODIES: ANTINUCLEAR AUTOANTIBODIES: *autoantibodies reacting with cell nuclear apparatus *present in patients with systemic disease of conective tissues *their level is associated with intensity of inflammation

  10. mitotic apparatus centromera ANA ENA: extractable nuclear antigens

  11. ANTINUCLEAR AUTOANTIBODIES systemic lupus erythematodes (SLE) dsDNA 90 % patients ssDNA 70-95 % patients Sm 20-40 % patients dsDNA+ Sm pathognomic for SLE SSA/Ro 20-60 % patients histons 95% patients (drug-induced SLE) Mixed Conective Tissue Disease (MCTD) U-RNP 95-100 % patients

  12. sclerodermy Scl-70 (DNA topoisomerasa-1) 50-70% patients CREST syndrom centromers 80-95 % patients polymyositis, dermatomyositis antisyntetase syndrom Jo-1 (histidyl-tRNA syntetáza) 25-30 % patients Sjögren´s syndrom (SS) SSA/Ro 40-95 % patients SSB/La 45-95 % patients

  13. DETECTION OF AUTOANTIBODIES BY INDIRECT IMMUNOFLUORESCENCE POSITIVE RESULT (AUTO Ab+) NEGATIVE RESULT (AUTO Ab-) LIGHT EMISSION (FITC  GREEN) UV LIGHT UV LIGHT NO EMISSION FLUOROCHROME - CONJUNG. ANTISERUM AGAINST HUMAN Ig WASHING WASHING NO Y SERUM (AUTO Ab) WASHING Y AUTO Ag NO WASHING CELLS GLASS

  14. HOMOGENNOUS NUCLEAR antigen:DNA, histons clinical association:systemic lupus erythematodes, rheumatoid arthritis, systemic sclerosis

  15. NUCLEAR MATRIX (large speckled) antigen: heterogenous ribonucleoproteins (hnRNP) clinical association: mixed connective tissue disease, systemic lupus

  16. FINE SPECKLEDFLUORESCENCE antigen:nuclear proteins (SS-A/Ro, SS-B/La, RNA polymerase II a III) clinical association:SLE Sjőgren´s syndrome, sclerodermy

  17. CENTROMERIC FLUORESCENCE (40-60 dots) Cells in telophase (metaphase): dots are in an equatorial area. antigen:kinetochors of chromosomes, centromeric antigens (CENP-A,B,C,D,E) clinical association:CREST syndrome, progressive systemic sclerosis

  18. NUCLEOLAR FLUORESCENCE antigen:PM-Scl complex of ten polypeptides similar to Ku antigen (66 a 68 kDa) clinical association:overlap syndrome

  19. dsDNA (native DNA) Substrate is protozoon Crithidia luciliae, with kinetoplast formed by dsDNA. . antigen: native dsDNA clinical association:significantly (95%) associated with SLE associated correlated with disease activity

  20. PRECISE DETERMINATION OF NUCLEAR FACTORS * molecularly characterized antigens * targets are produced by biotechnology * ELISA tests * immunobloting

  21. METHODS immunoblot ELISA recombinant or purified antigens

  22. ANCA ANTIBODIES * ANCA= antineutrophil cytoplasma antigens * substrate are ethanole fixed human granulocytes * present in certain types of vasculitis

  23. AUTOANTIBODIES ANCA AND VASCULITIS proteinase-3 myeloperoxidase

  24. cANCA antigen:proteinase 3 in  primary granules clinical association: Wegener granulomatosis

  25. pANCA antigen: myeloperoxidase (MPO). clinical association:cystic fibrosis (90%), chronic hepatitis (80%), primary sclerotiziing cholangoitis, SLE, rheumatoid arthritis

  26. ORGAN SPECIFIC AUTOANTIBODIES

  27. AUTOANTIBODIES AGAINST PARIETAL CELLS (APCA) antigen:antigen H+/K+/ATP-asa clinical association:pernicious anemia (80-90%), autoimmune endocrine diseases,

  28. LKM-1 antigen:cytochrome monooxygenase P-450IID6 clinical association:marker of autoimmune hepatitis

  29. AUTOANTIBODIES AGAINST RETICULIN (R1) antigen:reticulin R1 clinical association:celiakia

  30. AUTOANTIBODIES AGAINST ENDOMYSIUM (EMA) antigen:tissue transglutaminase clinical association:celiakia

  31. AUTOANTIBODIES AGAINST SMOOTH MUSCLE (ASMA) antigen:cytoskeletal antigens clinical asociation:marker of autoimmune hepatitis I, inflammatory diseases

  32. SUMMARY: THE PRESENCE OF AUTOANTIBODIES HAS TO BE INTERPRETED IN CLINICAL CONTEXT

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