This document discusses automated hematology analyzers and their use in analyzing complete blood counts. It describes 3-part analyzers, which provide white blood cell and platelet counts but have limitations, and 5-part analyzers, which provide more detailed analysis including a scatter plot. The document outlines the parameters measured by analyzers and how to interpret results, including normal ranges for red blood cell, white blood cell, and platelet counts and indices. Various abnormal findings are presented as case examples.

1. Dr Imtiaz Ahmed
MD Pathology, AFMC

2. Automated Hematology Analyzers
- Haemogram- Backbone of any lab evaluation
- As a routine investigation including anaemia,
polycythemia, infection, inflammation, allergy, drug
toxicity, malignancy, bleeding tendency etc
- Aim- to study RBC, WBC series and platelets

3. Sir Wallace H Coulter

5. 3 part analyser 5 part analyser
• Advantage: Cost effective
• Disadvantage:
1. Granulocytes taken together
2. Blasts counted as
lymphocytes
3. Cannot distinguish activated
lymphocytes and monocytes
4. Platelet count irregularities
5. No scatter plot
• Advantage:
1. Full differential
2. Platelet count more accurate
3. Flag for blasts, left shift,
giant platelets
4. Scatter plot obtained
5. Malaria parasite
• Disadvantage
1. Blasts still not detected
2. Newer parameters not given
3. Cannot distinguish between
activated lymphocytes and
monocytes/basophils

6. 3 Part Analyzer
Normal Report

7. 5 Part Analyzer

8. 5 Part Analyzer
RBC
WBC SCATTERGRAM
WBCPLT

9. Discrimination Thresholds
 WBC Discriminator
WBC LOWER discriminator-the optimum position in 30 - 60 fL .
Upper discriminator : 300 fL.
 RBC Discriminator
RBC LOWER discriminator- optimum position in 25 -75 fL and UPPER
discriminator, 200 - 250 fL.
RBC is calculated from the particle counts between this LOWER
discriminator and UPPER discriminator.
 PLATELET Discriminator
PLT LOWER discriminator, the optimum position in 2 – 6 fL and and
UPPER discriminator- 12 - 30 fL,

10. Sample intro
Hb
measurement
WBC count
& diff
RBC and
platelet count

11. Electrical Impedance
 Coulter principle
 Diluent displacement causes
potential difference.
 No. of impulse = No. of cells.
 Height = vol. of cells.
 Freq dist curve & size distribution
histograms
 Requisite – High dilution

12. Optical Light Scatter
 Each cell flows in a single line
through a flow cell
 A laser device focused
 On striking the cells scattering
in different directions
 Sensor capture
 Forward angle light scatter
(FALS)-Cell Size
 Side scatter(SS)-Granularity

13. Parameters Measured
Directly Measured Derived From Histograms Calculated
1.RBC Count
2.WBC Count
3.Platelet count
4.Hemoglobin
5.Reticulocyte
Count
1.MCV
2.RDW
3.DLC
4.PDW
1.Hematorit
2.MCH
3.MCHC

14. INTERPRETATION

15. Red Cell Indices

16.  Normal RBC count:
 Men: 4.5 to 5.5 x 106 per cumm
 Women: 4 to 5 x 106 per cumm

18.  Hemoglobin:
 Men: 13 to 16.5 g%
 Women: 12 to 16 g%
 Pregnant women: 11 to 16 g%

20. Mean corpuscular volume
 Normal: 80 to 100 fL
 Low MCV: Microcytosis
 High MCV: Macrocytosis
 Megaloblastic
 Non-megaloblastic

22. MCH/MCHC
 MCH: The amount, or the mass, of hemoglobin
present in one RBC, picogram
 26-32 pg
 MCHC: The amount of Hb in the hematocrit,
percentage
 32-37 g/dL

23. RDW
 Uniformity in size of RBCs
 11.5 - 14.5 %
 Low value: more uniform (<14)
 High values:
 Mixed population of small and large RBCs
 Anisopoikilocytosis

24. Reticulocyte count

25. Reticulocyte Production Index
Correction of reticulocyte count for the -
• degree of anemia
• duration of maturation

26. RBC Histogram
 Gaussian (Bell Shaped) curve
 Peak ideally within 80-100 fl
 2 flexible discriminator
LD (25-75fl)
UD(200-250fl)

27. RU-Flag
 Normoblasts
 Cold agglutinins
 Rouleaux formation
 ALL- L1
 Large platelets
 Fragmented RBCs
 Platelet
aggregation
RL- Flag

28. MP-Flag
 Blood transfusion
 Dimorphic anemia
 Treated IDA

29. White Blood Cells

30.  Normal range: 4 – 11 x 103 / cumm
 DLC:
 Neutrophils : 40 – 70% (2 - 8/cumm)
 Lymphocytes : 20 – 40 %. (1 - 3/cumm)
 Monocytes : 2 – 10 %. (0.2 – 1/cumm)
 Eosinophils : 1 – 6 %. (0.02 – 0.5/cumm)
 Basophils : 0 – 2 % (0.02 – 0.1/cumm)
• Immature cells
• Blasts
• Myelocytes
• Promyelocytes
• Metamyelocytes
• Band forms
• Activated lymphocytes

31. Leukopenia
 Acute viral infections
 Chemo/Radiation
 Aplastic anemia
 Collagen vascular diseases
 Bacterial Inf (Typhoid, Tuberculosis..)
 Parasitic infection (Malaria..)
 HIV
 Sepsis
 Drugs: immunosuppressants
Leukocytosis
• Infection (Bacterial)
• Inflammation
• Trauma, burns
• Leukemias

32. Neutrophils
 Absolute neutrophil count:
 Neutrophils x WBC / 100
 Normal : 2000 to 8000 / cumm
 Neutropenia:
 ANC < 1500/cumm

33. Neutropenia Neutrophilia
1. Decreased production
1. Aplastic anemia
2. B12, folate def
3. Severe sepsis
4. Leukemia
2. Increased destruction
1. Autoimmune
3. Marginalization and
sequestration
4. Chemotherapy
5. Viral infections
1. Acute infections
2. Steroids
3. Post splenectomy
4. Leukemoid reactions
5. Myeloproliferative
disorders

35. Lymphocytopenia Lymphocytosis
1. Infections
1. Typhoid
2. HIV
3. Viral hepatitis
2. Autoimmune disorders
1. SLE
2. RA
3. Steroids
4. Aplastic anemia
1. Acute viral infections
1. CMV
2. EBV
2. Chronic inflammatory
diseases
3. Tuberculosis
4. Stress
5. Leukemia/ lymphoma

36. Monocytopenia Monocytosis
1. Bone marrow failure
2. Hairy cell leukemia
1. Chronic infections
2. Malaria
3. Bacteria endocarditis
4. Collagen vascular diseases
5. Recovery from leukemia
6. CMML
7. M4/M5

37. Eosinophilia
1. Asthma
2. Other allergies
3. Parasitic infections
4. Inflammatory disorders (Coeliac disease, IBD)
5. Hypereosinophilic syndrome
6. Leukemias

38. Basophilia
1. Allergic reactions (Hay fever)
2. Inflammation (IBD, RA)
3. Tuberculosis
4. Diabetes Mellitus
5. Down’s syndrome
6. Neoplastic conditions
1. CML
2. PMF
3. ET
4. MDS

39. WBC HISTOGRAM
* Different in the three part analyser of our lab

40. WL-Flag
 Platelet aggregation
 Lyse resistant RBCs
 Cold agglutinin disease
 nRBCs.
 Giant platelets
WU- Flag
 Curve does not end at
baseline
 Immature WBCs
 Hyperleucocytosis

41.  Peak Between T1- T2: Acute Leukemia
 Peak Between LD- T1: CLL
 Peak Between T2- UD : Neutrophilia

43. Platelet
 Normal range: 150 – 400 x 103 / cumm
Thrombocytopenia Thrombocytosis
1. ITP
2. Splenomegaly
3. HUS, TTP
4. HELLP syndrome
5. Dengue
6. Drug induced
7. Transfusion associated
8. Aplastic anemia
9. Leukemia/Lymphoma
1. Reactive:
1. Chronic infection
2. Acute inflammation
3. Iron def
4. Post splenectomy
2. Myeloproliferative
disorders
3. Malignancies of lung, GIT

44. Platelet Histograms
 Between 2 discriminators
 Touch baseline
 LD (2-6fl)
 UD (12-30fl)

46. PL-Flag
 Cell Fragments
 Contamination-Bacteria
 Platelet aggregation
PU-Flag
 Clotted Blood
 Fragmented RBCs
 Microerythrocytosis
 Large platelets
 Platelet clumps

47. MP-Flag
 Platelet Anisocytosis
 Aggregation
 Recovery after chemo

48. Hemogram
Hb 9.5 g/dL TLC 4900/cumm
TRBC 3.2 x 106 DLC WNL
MCV 87 fL Platelet: 196,000/cumm
MCH 29 pg MPV 8.4 fL
MCHC 34 g/dL
RDW 17 %
Normocytic Normochromic Anemia

49. Microcytic hypochromic Anemia
Hemogram
Hb 8.4 g/dL TLC 4900/cumm
TRBC 2.8 x 106 DLC WNL
MCV 71 fL Platelet: 450,000/cumm
MCH 23 pg MPV 10 fL
MCHC 29 g/dL
RDW 21 %

50. Megaloblastic Anemia

51. Macrocytic Anemia
Hemogram
Hb 8.9 g/dL TLC 4100/cumm
TRBC 2.9 x 106 DLC WNL
MCV 107 fL Platelet: 154,000/cumm
MCH 29 pg MPV 10 fL
MCHC 35 g/dL
RDW 21 %

52. Hemogram
Hb 7.1 g/dL TLC 2100/cumm
TRBC 2.5 x 106 DLC N65 L28 M05 E02
MCV 118 fL Platelet: 72,000/cumm
MCH 31pg MPV 10 fL
MCHC 37g/dL
RDW 22.4 %
Aplastic Anemia/ Megaloblastic Anemia/
MDS

54. Marked macrocytosis with markedly increased MCH, MCHC
COLD AGGLUTININ DISEASE
Hemogram
Hb 5.6 g/dL TLC 17600/cumm
TRBC 1.96 x 106 DLC N89 L9 M1 E1
MCV 138 fL Platelet: 23,000/cumm
MCH 53 pg MPV 9.7 fL
MCHC 59 g/dL
RDW 26 %

56. Microcytic hypochromic blood picture
Hemoglobinopathies – (Beta – Thal trait, HbE
heterozygous)
Mentzer Index: MCV/RBC count, <13>
Mentzer Index: 9.6
Hemogram
Hb 12 g/dL TLC 7600/cumm
TRBC 5.8 x 106 DLC WNL
MCV 56 fL Platelet: 168,000/cumm
MCH 23 pg MPV 9.6 fL
MCHC 26 g/dL
RDW 21.8 %

57. Microcytic hypochromic blood picture
Hemoglobinopathies ( Beta thal major, HbE
heterozygous, HbE homozygous)
Mentzer Index: MCV/RBC count, <13>
Mentzer Index: >13
Hemogram
Hb 7.9 g/dL TLC 7600/cumm
TRBC 2.4 x 106 DLC WNL
MCV 56 fL Platelet: 168,000/cumm
MCH 23 pg MPV 9.6 fL
MCHC 26 g/dL
RDW 21.8 %

58. Hereditary Spherocytosis

60. Hemogram
Hb 13.2 g/dL TLC 7600/cumm
TRBC 4.31 x 106 DLC WNL
MCV 88 fL Platelet: 42,000/cumm
MCH 28 pg MPV 11.7 fL
MCHC 33 g/dL
RDW 13.2 %
Immune Thrombocytopenia
Bone marrow – Adequate megakaryocytes

61. Dengue Fever
Monocytosis/ Basophilia
Activated lymphocytes

62. Hemogram
Hb 8.9 g/dL TLC 17600/cumm
TRBC 2.8 x 106 DLC N89 L9 M1 E1
MCV 77 fL Platelet: 23,000/cumm
MCH 22 pg MPV 9.7 fL
MCHC 24 g/dL
RDW 23.1 %
3rd trimester pregnancy
HELLP syndrome

64. Hemogram
Hb 15 g/dL TLC 47600/cumm
TRBC 5.1 x 106 DLC N45 L50 M2 E3
MCV 109 fL nRBC 200/100 WBC
MCH 31.5 pg Corrected
TLC
15,600/cumm
MCHC 36.3 g/dL Platelet: 293,000/cumm
RDW 14.9 % MPV 9.7 fL
5 day child, Rh negative mother
Erythroblastosis foetalis

66. Hemogram
Hb 21 g/dL TLC 18600/cumm
TRBC 6.5 x 106 DLC N45 L50 M2 E3
MCV 91 fL Platelet: 593,000/cumm
MCH 31.5 pg MPV 9.7 fL
MCHC 36.3 g/dL
RDW 14.9 %
Panmyelosis
Polycythemia
70/M

67. Hemogram
Hb 9.1 g/dL TLC 108600/cumm
TRBC 2.9 x 106 DLC N75L7M2B16
MCV 91 fL Left Shift Marked
MCH 31.5 pg Platelet: 60,000/cumm
MCHC 36.3 g/dL MPV 9.7 fL
RDW 14.9 %
Chronic Myeloid Leukemia
35/M

68. Hemogram
Hb 9.9 g/dL TLC 50600/cumm
TRBC 3.1 x 106 DLC N75L7M2B16
MCV 81 fL Left Shift Marked
MCH 31.5 pg Platelet: 190,000/cumm
MCHC 36.3 g/dL MPV 9.7 fL
RDW 14.9 %
Leukamoid reaction
65/M, Uncontrolled DM, Fever, Cellulitis

69. Hemogram
Hb 10.1 g/dL TLC 108600/cumm
TRBC 2.9 x 106 DLC N5L92M2E1
MCV 91 fL Platelet: 60,000/cumm
MCH 31.5 pg MPV 9.7 fL
MCHC 36.3 g/dL
RDW 14.9 %
Chronic Lymphocytic Leukemia
65/M

70. Hemogram
Hb 10.1 g/dL TLC 108600/cumm
TRBC 2.9 x 106 DLC N5L92M2E1
MCV 91 fL Platelet: 40,000/cumm
MCH 31.5 pg MPV 9.7 fL
MCHC 36.3 g/dL
RDW 14.9 %
Acute Lymphoblastic Leukemia
4 yr/ M

71. WBC Research Population Data Case Study – Malaria Parasites(Normal plot and
Research Population Data compared to a patient infected with malaria type
Plasmodium falciparum. Note the increased size and variation of the lymph's and
Monocyte's.)
NORMAL
Normal
Normal MO
Normal LY
Macrophage
Parasitized
RBC
MALARIA
MP Positive
Reactive LY

72. INTERFERENCE
Hemoglobin Clotted specimen (micro-
clots).
Cryoglobulin, Haemolysis,
Severe lipemia,
Hyperbilirubinemia.
RBC count Clotting, Cold agglutinins,
Microcytic RBCs
Cryoglobulin, Giant platelets,
Raised WBC count.
WBC count Clotting, Cryoglobulin, Heparin,
Monoclonal proteins.
Platelets Clotting, Giant plt, Plt
clumps
Cryoglobulin, Haemolysis,
Microcytic RBCs.
Parameter Spurious Decrease Spurious Increase

73. Its all about
interpretation!!!