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CLS 3311 Advanced Clinical Immunohematology

CLS 3311 Advanced Clinical Immunohematology. Other Blood Groups II Ii, Le, P, Lu, Ch/Rg, Di, . Ii Blood Group GENETICS AND BIOCHEMISTRY. Genetics: Unclear

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CLS 3311 Advanced Clinical Immunohematology

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  1. CLS 3311Advanced Clinical Immunohematology Other Blood Groups II Ii, Le, P, Lu, Ch/Rg, Di,

  2. Ii Blood GroupGENETICS AND BIOCHEMISTRY • Genetics: Unclear • I and i genes: Theory is that their products aretransferase enzymes that attach repeating units of Gal and GlcNAc to the ABO Precursor Substance. Big I gene codes for branching of the PS. • The I and i antigen sites are considered uncompleted ABH active chains. Harmening,Page 176, Figure 8.5 • What blood group has the most I antigen sites?

  3. Ii Antigens Little iantigen is LINEAR • Found on cord cells, predominantly Big Iantigen is BRANCHED • Gradually convert from i to I during the first 18 months of life. Not all i converted to I, some i still present on adult cells, normally. • Analogy: I think of trees when I think of the transition from i to I. i is linear (straight like a baby tree) on newborn red cells while I is branched (like a mature tree) on adult red cells.

  4. Big I antigens Little i antigens

  5. Ii Antigens • Antigens Found in saliva, human milk, on leukocytes and platelets, in amniotic fluid, hydatid cyst fluid and urine. • Enhanced by enzyme treatment • High Frequency antigens: 99.9% • Dosage?Doesn’t make much sense, does it? • Adult i phenotype: i Ag persists into adulthood without conversion to I (Rare)

  6. Ii Antibodies: Anti-I • Non-RBC Immune • Common autoantibody. Cold reacting IgM. • Reacts with virtually all adult red blood cells • What is reactivity with cord cells? • Everyone has it. Why do we use only Monospecific IgG AHG today? You can thank auto anti-I for that. • Reactivity enhanced with enzyme treated cells • Benign Autoanti-I • Reacts at room temperature and can be picked up at AHG phase using polyspecific AHG reagent. • Pre-warm technique will remove or reduce reactivity • Cold technique will enhance weak reactions

  7. Ii Antibodies Pathologic Anti-I • Wide thermal range of reactivity and higher titer • Reacts at higher temperatures • Good Complement binding antibody • Implicated in Cold Hemagglutinin disease secondary to Mycoplasma pnuemoniaeinfections • Can maskclinically significant allo-antibodies • Auto Adsorption may be necessary to remove reactivity Anti-IH • Found in the serum of A1 phenotype - reacts strongly with RBCs high in H as well as I antigens.

  8. Ii Antibodies Antii • Reacts preferably with cord cells • Non-RBC immune - IgM • Reacts best at 4oC but is seen at room temperature. • Potent auto-anti-i found secondary to Infectious Mononucleosis. • Transient: Only present with active disease

  9. Lewis System:Genetics • Two alleles: Le and le codominant • Le gene codes for transferase enzyme • legene is a silent allele (amorph) • The Lewis antigens are NOT intrinsic to red cell membrane. • They are produced in the plasma (Secretions) and adsorbed from the plasmaonto the red cell membranes.

  10. Lewis Phenotype Frequencies

  11. Lewis System:Genetics • Le gene, similar to the ABO system, codes for the production of an enzyme. • Enzyme adds Fucose to the sub terminal GlcNAc on (ABH) Precursor Subst. to form LeaAg. This occurs ONLY in SECRETIONS. LeaAntigen This attachment can occur ONLY on a Type 1 Chain. Why?

  12. Type 1 Chain:#1 carbon of Gal is attached to the #3 carbon of GlcNAc. Lewis transferase enzyme adds Fucose in an (14) linkage to make Lea antigen. It can add this fucose ONLY to the Type 1 chain. Why? Type 2 Chain:#1 carbon of Gal is attached to the #4 carbon of GlcNAc.

  13. Le gene enzyme adds Fucose here: Following the action of the Le gene the Se gene enzyme now adds a fucose to the same PS in the SECRETIONS resulting in the formation of the Leb antigen. Leb Antigen What Genes are necessary for the production of Leb Ag’s? Le and Se genes. Remember, the Se gene codes for the presence of the H antigen in secretions!

  14. Lea Lea What Le antigens would be present in the secretions and on the RBC if you inherit the following genes? Lea Lea Lea Lea Lea Lea Lea Lea Lele and se/se Lea Lea Lea The Lewis red cell phenotype is: Le (a+ b–) Have Lea in secretions. Lea Lea Lea Lea Lea Lea Lea

  15. Leb Leb What Le antigens would be present in the secretions and on the RBC if you inherit the following genes? Leb Leb Leb Leb Lea Leb Leb Leb Leb Lele and Sese Leb Leb Lea The Lewis red cell phenotype would be Le (a– b+). Have Lea and Leb in secretions. Leb Lea Leb Leb Leb Leb

  16. Lewis System:Antigens • Presence of Lea antigens is dependant on the inheritance of the Le gene. • Presence of Leb antigens is dependant on the inheritance of the Le and Se genes. • Both are necessary to produce Leb antigens.

  17. Lewis System:Antigens • Phenotype frequencies: page 150, Table 7-1 • What is missing? • Where is Le (a+b+)? • Leb antigen preferentially adsorbs onto the red cell membrane over Lea. Lea is still in the plasma but not found on the red blood cells • Lewis antigens routinely adsorb onto and elute off of the RBC membrane. • Cord blood: Le (a-b-)

  18. Lewis System:Antigens Changes in Phenotype • Many pregnant woman LOSE their Lewis Antigens: Le (a-b-) during pregnancy. Common to see Lewis antibodies in obstetrical patients.Transient. Upon delivery the antibody fades and the previous Lewis phenotype returns. • Also seen in cancer patients, alcoholic cirrhosis, viral and parasitic infections.

  19. LewisAntibodies • Non-RBC Immune (IgM) • Good complement activators : can cause both in vivo and in vitro hemolysis • Enhanced reactivity with enzyme treated reagent red cells • Agglutination is weak, dirty looking. • Anti-Lea • Most common Lewis antibody encountered • Lewis antibodies are neutralized by Lewis substances.

  20. LewisAntibodies Anti-Leb Two categories of anti-Leb antibodies • Anti-LebH:Reacts best with Leb positive red cells with highest amount of H. Group O or A2 Leb positive red blood cells. More common of the two. • Anti-LebL:Reacts equally will with the Leb antigen on red cells of all ABO phenotypes.

  21. P Blood Group System • Genetics: These genes code for enzymes that sequentially add sugars to precursor substance. Sound familiar? It should. This system is related to the ABO, Le and Ii systems. • Genes: P1, P, Pk and lower case p (silent allele)

  22. P Blood Group System Biochemistry • All antigens are expressed in glycolipids on red cells, only, not in glycoproteins. • Antigens are built through the addition of sugars to precursor glycosphingolipids, analogous to A, B and H antigens. • P1 antigens are built by adding Gal in a (14) linkage to the same precursor substance (paragloboside) as H, A, B, I and i antigens are built.

  23. AABB Technical Manual, 14th Edition, Page 290.

  24. AABB Technical Manual, 14th Edition, Page 291.

  25. Deteriorates during storage Variable antigen expression between individuals Not well developed at birth Decreases in strength of expression during storage, therefore want to use fresh red cells to show the strongest expression of the P1 antigen Different reaction strengths between reagent red blood cells (Panel and Screen) No HDN P1Antigen

  26. P Blood Group System:Antibodies Anti-P1 • Found in P2 phenotype • Weak, cold reacting antibody • IgMclass antibodyreacts best at <25 oC • Non-RBC Immune (Naturally occurring) • Enhanced with enzyme treatment of reagent red blood cells: Screen and Panel cells.

  27. P Blood Group System:Antibodies Anti-P • Naturally occurring in Pk Phenotype • Auto Anti-P(IgG) found in Paroxysmal Cold Hemoglobinuria (PCH) • Called the Donath–Landsteiner antibody • Biphasic hemolysin: Binds complement at cold temperatures and activates that complement in warm temperatures lysing the red blood cells. • Found in children <14 y.o.following viral infection and tertiary syphilis

  28. Donath-Landsteiner Test • Draw two tubes of blood (clot tubes). • Tube 1: Incubate at 37oC for 60 minutes. (Control tube.) • Tube 2:Incubate at 4oC for 30 minutes transferring to 37oC for 30 more minutes. • Centrifuge and observe for hemolysis. • Positive: No hemolysis in tube 1 and hemolysis in tube 2. • Negative: No hemolysis is either tube. • Invalid: Hemolysis in tube 1.

  29. Very Clinically Significant Immunoglobulin class Non-RBC Immune Only found in amorph littlep phenotype HTR, HDN and implicated in early spontaneous abortions IgM with a IgG component. Wide thermal range, good C’ activator. Does NOT require foreign rbc stimulus Separable anti-P, anti-P1 and anti-Pk. Reacts with all cells except p phenotype P Blood Group System:Anti-PP1Pk (Tja)

  30. Lutheran Blood Group System • Genetics:Lua,Lub, Lu alleles • Codominant, Lu is silent allele Lua and Lub Antigens • Poorly developed at birth:MildHDN • Show Dosage • Variation of Ag expression between family members: Variable reactivity between cells on panel and screen. • Lu3 antigen: present when ever you have Lua or Lub antigens • Table 8-12 and 8-13 pages 189-190 Harmening

  31. Lu(a-b-) Phenotype • Recessive: Inherit two Lu genes (LuLu) • No detectable Lu antigens present • Make anti-Lu3 • In(Lu): Dominant Inhibitor (suppressor) inherited independently of Lutheran. • Carry trace amounts of Lu antigens • No anti-Lu3 antibody • X-Linked: Similar to In(Lu) • Trace amounts of Lu antigens present

  32. Lutheran Phenotype Frequency Lua = 7.65% & 5.3%Lub = 97.85% & 99.9%

  33. Lutheran Antibodies Anti-Lua • Non RBC Immune (Naturally Occurring) • IgG, IgM or IgA components present • React best at room temp • Most are clinically insignificant • See in crossmatch more likely than IAT • No HTR or HDN • Agglutination is loose mixed field

  34. Anti-Lub Anti-Lu3 RBC Immune, IgG class Reacts at 37oC and AHG phases Mild HDN and HTR Clinically significant 2.Inseparable anti-Lua and anti-Lub AHG phase, Reacts with all rbc’s except Lu (a-b-) Found in Lu (a-b-) phenotype Lutheran Antibodies

  35. Diego Blood Group System • Consists of two independent pairs of antigens: • Dia and Dib (Diego) • Wra and Wrb (Wright) • Dia/Dib • Dia antigen is almost entirely confined the population of Asian origin, including Native Americans. • One high (Dib) and one low (Dia) incidence antigen • RBC Immune: IgG class, activates complement • Causes HTR and HDN

  36. Diego Blood Group System • Wra low frequency Antigen (<0.1%) • Wrb high frequency Antigen (99.9%) Two types of Wra Antibodies • Non-RBC immune:IgM, IS, rxt’s better at 20oC than at 37oC • RBC-immune:IgG class, AHG phase, causes HTR an HDN, WAIHA • Wra is frequently encountered in the transfusion center due to immunogenicity of antigen

  37. Chido/Rogers Blood Group System • Chido and Rogers antigens are high incidence (95%) present on C4 component of Complement. • Antigens are NOT intrinsic to the red cell but are adsorbed on in the plasma. • Antibodies are generally benign but they are a “great nuisance in serological investigations” (TM).

  38. Chido/Rogers Antibodies High Titer, Low Avidity • High titer: 1:64 or greater • Low Avidity: 1+ or less through all titer dilutions • Agglutination is weak and variable with each cell on the panel. No pattern emerges. • RBC Immune: IgG, non Complement binding • Resolution: If you suspect an HTLA Ab you can do two things: 1) Titer patient serum: HTLA antibodies will titer out a long way (1:64) while others will not then you can… 2) Neutralize patient serum with normal fresh plasma, it will remove HTLA antibody.

  39. High Incidence Antigens: Table 9-3 Page 204 Vel • IgM with some IgG present • RBC Immune: HTR (rare), No HDN due to weak expression on cord cells • In vivo and In vitrohemolysis demonstrated Sda (Sid) • RBC Immune: IgM with few IgG • RT and AHG reactions enhanced with enzymes • MF agglutinates, shiny refractile agglutination • No HDN and HTR is very rare

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