1. LS/MW MT 417 – Clinical Hematology II Manual/Special Tests Unit Ham’s/Sugar Water/DL Exercise Ham’s/Sugar Water/DL Questions KEY

2. LS/MW 1.For what conditions are the Ham’s and Sugar water tests performed? Performed to detect Paroxysmal Nocturnal Hemoglobinura (PNH); refers to intermittent, hemoglobin in the urine, after night. Individuals with PNH have an acquired membrane defect in which the red cell membrane has an increased sensitivity for complement to bind than do normal red cells. Complement binding causes holes in the membrane = complement-induced RBC lysis. What is the defect in this condition?

3. LS/MW 1.Is it antibody mediated? NO antibody is involved and the cause of PNH is unknown. All cells are abnormally sensitive to lysis by complement. It is characterized by pancytopenia, chronic intravascular hemolysis that causes hemoglobinuria and hemosiderinuria. PNH is also noted for a low LAP score, problems with thrombosis & infection and an increased incidence of leukemia.

4. LS/MW 2.ABO compatible serum is the source of complement for both the Ham’s and Sugar water tests. Which test uses sucrose to enhance complement binding? The sugar water test uses a low-ionic strength sucrose solution. The Ham’s test uses acidified serum. 2.Which test uses acid to enhance complement binding?

5. LS/MW 2.What indicates a positive result? In both tests, the presence of hemolysis in the ‘test’ tube(s) is positive for the presence of PNH red cells that are sensitive to complement binding. The sucrose hemolysis test is the screening test; the Ham’s acid serum test is specific for PNH. PNH can also be diagnosed using immunophenotyping. Positive Sugar Water Test indicated by hemolysis in tube#1 Control tubes #2-4 negative for hemolysis

6. LS/MW 3.For what condition is the Donath-Landsteiner test performed? Performed to detect Paroxysmal Cold Hemoglobinuria; refers to intermittent, hemoglobin in urine, after cold. This disorder is antibody-mediated and may be idiopathic or associated with acute viral infection. It is characterized by intravascular hemolysis and hemoglobinuria after cold exposure due to the Donath-Landsteiner antibody that causes complement-induced red cell lysis. Is this condition antibody mediated?

7. LS/MW 4.In the Donath-Landsteiner test, tubes are observed for hemolysis following incubation in the cold and/or at 37°C. Why? Testing for the presence of a biphasic autoantibody that fixes complement to the red cells in the cold and when the temperature rises, the complement-coated red cells lyse. The DL hemolysin is an IgG antibody with P specificity that recognizes the RBC membrane as foreign in the cold.

8. LS/MW 5.A 59-year-old woman presents with blood and bone marrow findings of aplastic anemia. She gives a history of occasional morning episodes of dark urine. Which of the following tests would you expect to be abnormal? a.Direct antiglobulin test and osmotic fragility test c.Sucrose hemolysis test and Ham’s acid serum test Findings of pancytopenia and hemoglobinuria after night are suggestive of PNH....expect abnormal Ham’s and Sugar water tests. b.Cold agglutinin titer and Donath-Landsteiner test

9. LS/MW 6.Which of the following is characteristic of both PNH and PCH? b.Hemoglobinuria after night a.Complement-induced lysis c.Hemoglobinuria after cold d.Acquired membrane defect e.Autoimmune disorder a. Complement-induced lysis occurs in both PNH (b. and d.) and PCH (c.)

10. LS/MW You have completed the Ham’s/Sugar Water/DL Exercise! Positive Sugar Water Test indicated by hemolysis in tube#1 Control tubes #2-4 negative for hemolysis