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laboratorydiagnosisofanemiaugs-170319165804 (1).pptx
- 1. Laboratory Diagnosis ofAnemia By Dr.Varughese George Department of Pathology
- 2. Learning Objectives At theend of this briefing, you should know • Clinical presentation of anemia. • Definition of anemia. • Approach to diagnosis of anemia • Classification of anaemia • Distinct features of each type of anemia.
- 3. Clinical presentation ofanemias • Fatigue and weakness • Headache • Tinnitus • Numbness and coldness • Pallor • Dyspnea and palpitations • Angina pectoris • Intermittent claudication • Hemorrhages in the fundus of eyes • Menorrhagia • Anorexia • Flatulence • Nausea • Constipation
- 4. Clinical presentation ofanaemias
- 5. Initial Laboratory Work-Up •Hemoglobin concentration. • Packed Cell Volume. • Red cell indices. • Peripheral Blood Smear.
- 6. Definition of Anaemia •Anaemia is defined as a reduction in the concentration of circulating haemoglobin below the level that is expected for healthy personsof same age and sex in the same environment. 6
- 7. Blood parameters (LaboratoryNormal Range)
- 8. Blood parameters (LaboratoryNormal Range) •In full term infants, hemoglobin is 18.0 ± 4.0 g/dl •Children - 12.5 ± 1.5 g/dl •Children(6-12 years) - 13.5 ± 2.0 g/dl
- 9.
- 10. Classification of Anaemias •Morphological Classification • Etiological Classification • Classification based on reticulocyte response.
- 11.
- 12. Hematocrit • Proportion ofthe volume of red cells relative to the volume of blood • Rules of Three: – RBC X 3 = Hemoglobin – Hemoglobin X 3 = Hematocrit Packed cell volume (PCV) or Haematocrit (Hct) Men - 0.45 ± 0.05 l/l (40-50%) Women - 0.41 ± 0.05 l/l (38-45 % in non- pregnant women 36-42 % in pregnant women)
- 13. Mean Corpuscular Volume •Dividing the total volume of red cells by the number of red cells • Index for average size of red cells • Normal range - 92 ± 9 fl 13
- 14. Mean Corpuscular Hemoglobin •Average amount of haemoglobin in each red cell. • It is expressed in picograms or pg. • Normal range - 29.5 ± 2.5 pg
- 15. Mean Corpuscular HemoglobinConcentration • This represents the average concentration of haemoglobin in a given volume of packed red cells. • Normal range – 330 ± 15 g/l • MCHC raised in hereditary spherocytosis. • Decreased in hypochromic anaemia.
- 16. Red cell distributionwidth • Variation in red cell size • Normal range - 12.8% ± 1.2% • Low in B-thalassemia trait • High in iron deficiency anaemia • Normal in anaemia of chronic disease
- 17. 17 1 Microcytic/hypochromic 3 1 2 2Macrocytic/Normochromic 3 Normocytic/Normochromic Morphologic Categories of Anemia N.B. The nucleus of a small lymphocyte (shown by the arrow) is used as a reference to a normal red cell size
- 20. Macrocytic anemia • Low/normalreticulocyte count, macrocytosis(oval and round) • Elevated MCV,MCHC • Basophilic stippling • Howell-jolly bodies • Cabot rings • Pancytopenia • Hypersegmented neutrophils • Bone marrow- megaloblastic maturation,sieve like chromatin,nuclear- cytoplasmic asynchrony,maturation arrest
- 21. Normocytic Normochromic anaemia witheffective erythropoiesis
- 22. Etiological Classification ofAnaemias
- 23. Basic Approach toa diagnosis of anemia
- 24. Evaluation of microcytichypochromic anaemia
- 25. Evaluation of macrocyticanaemia
- 26. Evaluation of normocyticanaemia
- 27. Evaluation of haemolyticanaemia
- 28. A simplified approachto diagnosis of haemolytic anaemias
- 29. Reticulocyte Count (In theDiagnosis of Anemia) • Reticulocytes are non- nucleated RBCs that still contain RNA. • Visualized by staining with supravital dyes, including new methylene blue or brilliant cresyl blue. • Useful in determining response and potential of bone marrow. • Normal range is 0.5-2.5% of all erythrocytes. 29
- 30. Fe++ deficiencyanemia • Lowhemoglobin and low packed cell volume • Low MCV,MCH and MCHC • Microcytosis & hypochromia are hallmarks • RDW is increased • Serum ferritin is less than 15 micro gram/dl • Serum iron is low,TIBC is increased and transferrin saturation is less than 10 percent • Free erythrocyte protoporphyrin is increased. • Increased soluble transferrin receptor in serum • Bone marrow-micronormoblastic,absence of stainable iron in bone marrow on Perls Prussian blue reaction
- 31. 31 Megaloblastic Anemia Mild tosevere anemia, – Increased MCV & MCH, normal MCHC – Low RBC, HGB, WBC and PLT counts (fragile cells) due to ineffective hematopoiesis. – Low reticulocyte count – Macrocytic ovalocytes and teardrops; – Marked anisocytosis and poikilocytosis – Schistocytes/microcytes - due to RBC breakage upon leaving the BM – Erythroid hyperplasia - low M:E ratio (1:1) – Iron stores increased.
- 32. 32 Macrocytic Ovalocytes Blood NRBCBlood Howell-Jolly body Teardrop Schistocyte Stippled RBC & Cabot Ring Giant Platelet Pap bodies Hypersegmented Neutrophil >5 lobes Megaloblastic anemia
- 33. Tests • Folate andB12 levels • Schilling test may be useful to establish etiology of B12 deficiency – Assesses radioactive B12 absorption with and without exogenous IF • Other tests if pernicious anemia is suspected – Anti- parietal cell antibodies, anti-IF antibodies – Secondary causes of poor absorption should be sought (gastritis, ileal problems, etc.)
- 34. Anemia of chronicdisease • Normocytic anemia with ineffective erythropoiesis (reduced reticulocyte count) • Normochromic • Results from – Chronic inflammation (e.g. rheumatologic disease): Cytokines released by inflammatory cells cause macrophages to accumulate iron and not transfer it to plasma or developing red cells (iron block anemia) – Inflammation – malignancy • Bone marrow suppression (EPO is elevated)
- 35. Anemia of chronicdisease • Decreased serum iron,decreased total iron binding capacity and normal or raised ferritin • Increased marrow storage iron • ESR is high
- 36. Normochromic, normocytic anemiawith effective erythropoiesis INCREASED reticulocyte count • Acute blood loss – Very acutely, with hypovolemia, may have normal blood counts, will become anemic with volume replenishment • Hemolytic anemia – Increased reticulocyte production cannot keep pace with loss of RBCs peripherally. • Response to specific therapy in nutritional anemias
- 37. Aplastic anemia • Pancytopeniacaused by bone marrow failure…decreased production of all cell lines and replacement of marrow with fat. • Inherited- Fanconis anaemia, Dyskeratosis congenita • Acquired - Idiopathic,drugs like NSAIDs,chloramphenicol,benzene,parvo virus,hepatitis and EB virus.
- 38. Hemolytic anemia • Abnormalityintrinsic to red cells- 1. Hereditary spherocytosis 2. Thalassamia 3. Sickle cell anaemia 4. Glucose -6-phosphate dehydrogenase deficiency • Abnormality extrinsic to red cells- 1. Immune 2. Mechanical etc
- 39. Evaluation of haemolyticanaemia
- 40. Hereditary spherocytosis • Inheriteddefect in the red cell membrane cytoskeleton (spectrin, ankyrin or band 3) leading to the formation of spherocytic red cells. • Autosomal dominant • Mild to moderate anaemia • Intermittent jaundice • Splenomegaly • Pigment gall stones • Peripheral smear-microspherocytes • Screening test-osmotic fragility
- 41. Thalassemia • Decreased orabsent globin chains • Alpha and beta thalassemias • Microcytic hypochromic,target cells,basophilic stippling • Reticulocytosis • Hb F elevated in electrophoresis
- 42. Sickle cell anaemia •Presence of Hb S • Point mutation in 6th place of beta chain • Substitution of valine for glutamic acid • On deoxygenation,sickle cells are formed • Chronic hemolytic anaemia,vaso- occlusive crisis • Aplastic crisis • Hemolytic crisis • Infections
- 43. Sickle cell anaemia •Sickling test is positive. • Solubility test is positive. • Electrophoresis shows HbS. • In sickle cell trait, electrophoresis shows 60 percent of Hb A and 40 percent Hb S
- 44. Glucose-6-phosphate dehydrogenase deficiency • Xlinked disorder • Reduced activity of G6PD • Inability to remove H2O2 • Accumulated H2O2 leads to oxidation of hemoglobin with precipitation of globin chains • Heinz bodies • Red cells with heinz bodies destroyed in spleen(extravascular hemolysis)
- 45. Glucose-6-phosphate dehydrogenase deficiency • Asymptomatic •Neonatal jaundice • Acute hemolytic anaemia • Chronic hemolytic anaemia • On peripheral smear- polychromasia,fragmented red cells,spherocytes,bite cells,half ghost cells • Biochemical-increased bilirubin,hemoglobinemia and hemoglobinuria
- 46. Glucose-6-phosphate dehydrogenase deficiency • Screeningtests-fluorescent spot test,methemoglobin reduction test and dye decolorisation test
- 47. Immune hemolytic anaemia •Warm antibody-persons over 50 years,mild jaundice and splenomegaly,red cells coated with IgG,spherocytes.Seen in autoimmune disorders,lymphoma • Cold antibody-acrocyanosis,IgM.Seen in cold agglutinin disease, Paroxysmal cold hemoglobinuria (PCH)
- 48. Coomb’s test • Detectspresence of either antibody on RBC or of antibody in serum • Helpful in determining if a hemolytic anemia is immune-mediated
- 49. SUMMARY • Microcytic hypochromicanaemia-iron deficiency • Macrocytic hyperchromic-megaloblastic anaemia • Normochromic normocytic-hemolytic anaemia • Pancytopenia-megaloblastic and aplastic anemias