Peripheral smear

Peripheral smear

1. PERIPHERAL SMEAR

2. PERIPHERAL SMEAR The role Examination of a well-made peripheral blood smear is Estimate approximately the numbers of each of the three cellular elements Red blood cells White blood cells Platelets b)Study the morphology of these c)See blood parasites

3. RED CELL MORPHOLOGY a) SIZE A Normal red blood cell is about 7.2micrometer in diameter A normal red blood cell should be approximately the same size as a normal lymphocyte nucleus When the variation in size is greater than the normal , this is referred to as Anisocytosis

4. Normocytic RBC

5. Microcytes When red cells are smaller than normal ,having a diameter of less than 7.2micrometer and MCV < 80 fl they are called Microcytes When the significant number of the cells are smaller than normal cells are referred to as Microcytosis

6. Macrocytes Macrocytes are larger red cells with a diameter of >7.8µ and MCV > 100 fl The significant number of the cells are larger than normal cells are referred to as Macrocytosis

7. .. b) SHAPE Normal red cells are biconcave discs and appear circular on a smear When the variation in shape is greater than the normal this is referred to as Poikilocytosis

8. POIKILOCYTOSIS They larger variety of specific abnormal shapes are present including OVALOCYTES OR ELLIPTOCYTES PENCIL SHAPED CELLS SPHEROCYTES TARGET CELLS OR LEPTOCYTES SICKLE CELLS SCHISTOCYTES CRENATED CELLS ACANTHOCYTES BURR CELLS STOMATOCYTES etc

9. ACANTHOCYTES Irregularly spiculated Hypochromic RBCs CONDITIONS: -Alcoholic liver disease - Lipoprotineamia -Post splenectomy - pyruvate kinase deficiency

10. TARGET CELLS OR LEPTOCYTES Hb is redistributed Center and periphery of the cell appear haemoglobinised . CONDITIONS: -Thalassemia -Obstructive liver disease

11. DACROCYTE(TEAR DROP CELL Cells with single elongated or pointed extremity Shaped like a tear drop CONDITIONS : - Myelofibrosis -Severe haemolytic anaemia

12. SICKLE CELLS Thin,elongated,slightly curved red cells Sickle shaped No central pallor area CONDITIONS: -Sickle cell disorders

13. ECHINOCYTES(CRENATED CELL) Crenation is the shrinkage of red cells through loss of water in a hypertonic medium. Cell membrane appear wrinkled. CONDITIONS: -As an artifact due to slow drying of smear. -Seen in smears made several hours later after collection -Uremia

14. BURR CELLS Numerous , uniform sized and irregularly spaced red cell projections. CONDITIONS: -Uremia -Chronic liver disease - pyruvate kinase deficiency

15. ELLIPTOCYTES(OVALOCYTES) Elongated or oval cells CONDITIONS: -Hereditary elliptocytosis -Thalassemia -Iron deficiency

16. SPHEROCYTES Spherical red cells Lack of central pallor area 2types Macrosperocytes Microspherocytes CONDITIONS : -Hereditary spherocytosis -Auto immune haemolytic anaemia

17. STOMATOCYTE Central pallor area appear like a slit. CONDITIONS: -Hereditary stomatocytosis -Obstructive liver diseases

18. SCHISTOCYTE(fragmented cell) Irregular shaped triangular spiculated cells due to fragmentation. CONDITIONS: -DIC -Hemolytic anemia

19. KERATOCYTE(Helmet cell) Have 1 or 2 projections which may vary in length. CONDITIONS: -Hemolytic anemia -Intravascular coagulation -Iron deficiency anemia

20. RED CELL INCLUSIONS Red cell cytoplasm may contain various inclusions which pick up some stain and visible Basophilic stippling Howell –jolly bodies Cabot rings Pappenheimer bodies

21. Cabot rings Pale staining rings or figures of eight in the RBC These are probably artifacts resulting from damage to lipoprotein of the stroma of red cells Seen in Haemolytic anaemia,megaloblastic anaemia,leukaemia and after splenctomy

22. Basophilic stippling RBCs show presence of numerous basophilic granules distributed through the cell cytoplasm These inclusions are precipitated ribosomal RNA This is found in metal poisoning megaloblastic anemia Hemolytic anemia thalassemia

23. Howell –jolly bodies Small round densely stained dark blue particles 1µm in diameter in the periphery of red cells These are remnants of nucleus (aggregates of chromatin material ) Feuglen positive for DNA

24. Pappenheimer bodies These are aggregates of ferritin and are located close to cell membrane . During erythropoesis when hemoglobin synthesis is not complete some iron granules are left behind in the red cell forming siderotic granules or Pappen - heimer bodies

25. Pappenheimer bodies With Romanowsky stain these appear pale blue but easily demonstrable with Perl’s stain.such red cells with Pappenheimer bodies are called siderocytes Sideroblastic aneamias Megaloblastic anaemia Hemolytic anaemia Post splenectomy

26. COLOUR When RBC are stained with one of the Romanowsky stains,they take up pink colour Because of the biconcave shape the stain darker at the periphery and lighter at the centre ( ie , area of the central pallor)

27. NORMOCHROMIC RBC When RBC show this typical staining ,they are said to be Normochromic area of central pallor which may be up to a third of the diameter of the cell

28. Hypochromia When the area of the central pallor is enlarged (more than 1/3 of the diameter of RBC ) the cells appear more pale ,and are said to be Hypochromic

29. Polychromasia Cells may appear a little bluish colour amidst the red cells of the haemoglobin , so they appear a little purplish or greish , known as Polychromotophilic This staining is found in young erythrocytes ,just after their release from bone marrow,when they still contain RNA in the cytoplasm .RNA picks b lue colour , reticulocytes

30. RETICULOCYTES Reticulocytes are immature RBCs released from bone marrow These RBCs are slightly larger with 20% more volume than RBCs Reticulocytes are stained in a living state invitro using basic dyes like Brilliant cresyl blue and New methylene blue known as supravital stain ,demonstrating blue filament ous or granular material

31. Morphology of WBC Leukocytes or white blood cells are another formed elements of blood .There is usually 1 WBC for every 500 RBC present . WBCs are divided into two types Granulocyte : cytoplasm contain granules Agranulocyte : No granules in the cytoplasm There are three types of granulocyte named according to their staining characteristics in blood films. They are neutrophils , eosinophils and basophils . Mononuclear cells are divided into lymphocytes and monocytes .

32. Morphology of WBC

33. NEUTROPHIL Diameter: 12-16 µm pink/orange cytoplasm with fine granulation Nucleus: dark purple blue dense heterogeneous chromatin 2-5 lobes

34. EOSINOPHIL Diameter: 14-16 µm Cytoplasm : full of granules Granules: large retractile orange-red Nucleus: blue dense chromatin Normally 2 lobes

35. BASOPHIL Diameter: 14-16 µm Cytoplasm: pink Granules: dark blue – black obscure nucleus Nucleus: blue

36. LYMPHOCYTE Diameter: small 7-9 µm large 12-16 µm Cytoplasm : abundant clear, pale blue Granules: small(a granular) large ( a variable number of azurophilic pinkish-purple granules ) Nucleus: dark blue \round dense homogeneous chromatin

37. MONOCYTE Diameter: 14-20 µm ( are the largest normal blood cells) Cytoplasm: grey blue Granules: a very fine pinkish blue granules Nucleus: blue large irregularly shaped and folded (kidney shaped or horse shoe shaped)

38. … TC 4,000-11,000 cells/cu mm Neutrophil = 40-75% Lymphocyte = 20-40% DC Eosinophil = 1-6% Basophil = 0-1% Monocyte = 2-10%

39. Variations in the WBC count T otal WBC count Increase in Total WBC count > 11,000 /µl is known as Leukocytosis Decrease in Total WBC count < 4,000 / µl is known as Leukocytopoenia CORRECTION FOR NUCLEATED RBCs 10 or > NRBCs - Make correction uncorrected WBC count X 100 number of nRBC’s per 100 WBC’s + 100

40. Variations in Neutrophils Neutrophila ;- Increase in the percentage of neutrophils in the differential count .it may be Relative Neutrophila ;-the total count is lowered by decreased numbers of lymphocytes Absolute Neutrophila ;- the total number as well as the percentage of neutrophils Neutropoenia ;- Decrease in the percentage of neutrophils in the differential count

41. Variations in lymphocytes Lymphocytosis Relative Lymphocytosis ; Absolute number of lymphocytes in blood may not be increased ,but due to a decrease in the number of other cells esp. Neutrophils ,the percentage of lymphocytes in the DC is increased Absolute Lymphocytosis ; an actual increase in the total number of lymphocytes in the blood as well as the percentage of lymphocytes in the DC is increased Lymphopoenia ; Decrease in the percentage of lymphocytes in the differential count

42. Variations other white blood cells Monocytosis Eosinophilia Basophilia

43. Evaluate WBC morphology Note if any abnormal white cell morphology is present Toxic granulation Dohle bodies Cytoplasmic Vacuolation Hypersegmentation Hyposegmentation Smudge cells

44. Toxic granulation Neutrophil contain prominent granules CONDITIONS: -Severe bacterial infection -Hereditary disorders

45. Dohle bodies Presence of small round or oval light blue stained bodies in the periphery of the neutrophilic cytoplasm. CONDITIONS: -Severe infections -Burns -Exposure to toxic agents

46. Cytoplasmic Vacuolation Occurrence of vacuoles in the cytoplasm and nucleus,due to the reactive changes seen in infections CONDITIONS: -Severe infections -Burns -Chemical poisoning -Malignancy

47. Hyper segmentation More than 4 lobes in neutrophils . CONDITIONS: -May due to inherited disorders - macrocytic anemia

48. Hypo segmentation Failure of normal lobe development(<3 lobes). CONDITIONS: -Acute myelocytic leukemia -Severe infections -Toxic states -Hereditary disorders

49. LEFT SHIFT

50. SMUDGE CELLS Degenerated lymphocytes with out cell wall. CONDITIONS: -Presence of few-> faulty technique -Large amount-> chronic lymphocytic leukemia

51. PLATELETS Examine blood smear for thrombocytopenia – confirm counts, cause Platelet count – – about 10 – 20 per oil immersion field or number of platelets in 10 oil.imm. fields X 15,000 select area where RBCs barely touch Falsely low platelet ct – small clots, Platelet aggregates platelet satellitism , abnormally large platelets

52. PLATELETS Blood smear for high platelet counts – confirm counts, cause Spurious thrombocytosis – red cell fragments, fragments of leukaemic cells, fungi, bacteria NORMAL PLATELETS: 1.5 – 4 lakhs / cu mm

53. . ABNORMALITIES OF PLATELET GIANT PLATELET Larger plateles upto 7 micrometers in size

54. Platelet satellitism Platelet satellitism describes the phenomenon of adherence of platelets to white cells. It is an in vitro phenomenon of no clinical significance. However it is important that it is detected since the platelet count will be low.

55. Platelet aggregates Platelet aggregates may becomposed of apparently intact platelets, degranulated pale grey platelets or a mixture of both, as in this example. If the platelet count is low it is essential to examine the blood film carefully for platelet aggregates.

56. HAEMOPARASITES Examine carefully for the presence of of any haemoparasite Malaria Microfilaria Trypanosoma LD bodies in monocytes Borrelia in relapsing fever Leptospira inWeils disease

57. HAEMOPARASITES Malarial Parasite Microfilaria

58. .. Trypanosoma LD bodies in monocytes

59. .. Borrelia in relapsing fever .

60. REPORTING FORMAT Normal blood picture RBC: Normocytic and Normochromic WBC: Count and distribution within normal limits Platelets : seen adequate Haemoparasite : Not seen Impressioin : Normocytic Normochromic blood picture.

61. References CMC manual A Practical manual -clinical Pathology, Sabitri Sanyal & AparnaBhattacharyya 2 Edition Clinical Pathology; Thejendra singh Dacie and Lewis: PRACTICAL HAEMATOLOGY

62. .. Thank you