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1. Haematology laboratory test result
interpretation and an approach to
anaemia diagnosis
Dr. Andy Hughes
Consultant Haematologist

2. Case study 1
• 25 year old female Cypriot.
• c/o tiredness -> FBC
• Hb = 10.5
• MCV = 65 (82-96)
• RBC = 5.5 (3.9-5.0)
• MCHC = 32 (32-35)
• RDW = 12 (10-14)

3. Case study 1
• Microcytic anaemia
• Possible diagnoses?
• Iron deficiency
• Thalassaemia trait
• Additional tests?
• Ferritin – 50; iron deficiency unlikely
• Hb A2 – 5% - raised
• Confirms β thalassaemia trait

4. Case study 2
• 70 year old Caucasian male
• c/o recurrent mouth ulcers
• Hb = 10
• MCV = 105
• WCC = 2.1 (>4.0)
• Neutros = 0.5 (>2.0)
• Platelets = 120 (>150)

5. Case study 2
• Macrocytic anaemia with a pancytopenia
• Further investigations?
• B12 = 180 (>240)
• Serum folate = 5.5
• LFTs - normal
• TSH - normal
• Retics = 25 (<1%)

6. Case study 2
• Course of B12 injections
• Hb 10.2
• Referred for bone
marrow
• Myelodysplasia
Pelger neutrophil
Bone marrow

7. Anaemia Classification
Morphology
Microcytic Macrocytic Normocytic

8. The FBC
• Hb
– anaemia
• Hct/PCV
– polcythaemia
• RBC
– iron deficiency and
thalassaemia trait
• RBC indices
• MCV
– Anaemia classification
• MCH/MCHC/RDW
– Differentiation of
microcytic anaemias

9. Red cell distribution width - RDW
• Measure of RBC size
variation (anisocytosis)
• Increased in iron
deficiency
• Normal in thalassaemia
trait
• Normal/raised in anaemia
of ch. Disease
• Can be very high in
megaloblastic anaemias
• ?practical value

10. Microcytic anaemia
• Decreased ferritin
• Decreased iron and raised TIBC
• Can be falsely raised with active chronic disease (eg RA)
Iron deficiency
• Normal ferritin
• Raised HbA2 (β but not α thal trait)
Thalassaemia
trait
• Normal/raised ferritin
• Decreased iron and TIBC
• Raised ESR/CRP
Anaemia of
chronic disease

11. Iron deficiency/Thal trait
• Iron deficiency
• Decreased Hb and RBC
• Decreased MCV+
• Decreased MCHC
• Increased RDW
• Thrombocytosis
• Film – pencil cells
• Thalassaemia trait
• Normal/dec. Hb and
normal/inc. RBC
• Decreased MCV++
• Normal MCHC
• Normal RDW
• Normal platelet count
• Film – target cells

12. Iron deficiency/thal trait
Blood films
IRON DEFICIENCY THALASSAEMIA TRAIT

13. Iron Deficiency
• Always confirm with
ferritin
• May be falsely normal
with active
acute/chronic disease
• If >50 makes iron
deficiency less likely
• In early deficiency or
with mixed deficiencies
MCV may be normal
• Causes
• Diet
– Young and old
• Malabsorption
– Coeliac (tTG)
• Chronic blood loss
– GIT
– Uterine
– UGT

14. Failure to respond to iron
• Inadequate treatment
– Dose and/or duration
– Avoid slow release preparations
• Continued cause
– Bleeding
• Malabsorption
• Poor compliance
• Intolerance due to side effects
• Incorrect diagnosis

15. Parenteral iron
• Indications:
• Genuine intolerance
• Malabsorption
• Gastric/upper GI surgery
• Inflammatory bowel disease
• IM
– CosmoFer
– Z-track technique
• IV
– Venofer as repeated small
volume injections
– CosmoFer as TDI
• Problem with
allergy/anaphylaxis
• Avoid if h/o
– Drug reaction(s)
– Allergy
– Asthma

16. Increased Ferritin
• Infection/inflammation
• Chronic liver disease; esp. alcohol related
• Malignancy
• Repeated transfusions
• Hereditary haemochromatosis
– >1000 is risk of organ damage
• Porphyria Cutanea Tarda
• Prolonged iron therapy without iron
deficiency

17. Case study 3
• 65 year old Caucasian male.
• Known stable, mild CLL; no treatment.
• c/o increasing fatigue and breathlessness.
• o/e jaundiced.
• No enlarged lymph nodes or spleen.

18. Case study 3
• Initial tests
• Hb = 8.
• MCV = 102.
• WCC = 30 (prev. 15).
• Lymphs = 25 (prev. 12).
• Platelets = 300.
• Bilirubin = 60 (<20).
• Other LFTs normal.
• Additional tests
• Retics 300.
• LDH 900 (<450)
• Conjugated bilirubin =
10.
• Direct antiglobulin
(Coombs) test DAT} –
positive.

19. Case study 3
• Macrocytic anaemia.
• Macrocytosis due to increased reticulocytes
due to secondary haemolysis.
• Haemolysis caused the jaundice.
• Haemolysis autoimmune (positive DAT)
• Diagnosis:
– Autoimmune haemolysis associated with known
CLL.
• Treatment steroids.

20. Tests for haemolysis
• Initial evaluation
• LFTs
– Inc. bilirubin only
• Retics - increased
• LDH - increased
• Haptoglobin -
decreased
• Additional tests
• Blood film
– Polychromasia
– Spherocytes
– Sickle cells
• Hb electrophoresis
• Direct Antiglobulin
(Coombs) test

21. Haemolysis –blood films
Sickle cell Spherocytes

22. Macrocytosis
• B12/folate deficiency
• Chronic liver disease
• Excess alcohol
• Myelodysplasia
Common
• Haemolysis
• Myeloma
• Other bone marrow disorders
• Hypothroidism
Less
common

23. Blood films in macrocytosis
Target cells in liver disease Polychromasia

24. B12 and folate deficiency Megaloblastic
anaemia
Hypersegmented polymorphs
Oval macrocytes and
poikilocytes

25. Megaloblastic bone marrow

26. Megaloblastic anaemias
• Dec. Hb with raised MCV (often >115).
• Inc. RDW; can proceed a rise in the MCV.
• With severe anaemia  inc. in poikilocytosis
 dec. in MCV and a rise in RDW ++.
• Other cytopenias.
• Film – hypersemented polymorphs.
• Very high LDH.
• Slightly increased bilirubin.

27. Vitamin B12 deficiency
• B12 Levels can vary; may need to repeat.
• Check diet; especially vegans.
• MCV may be helpful if >115
• Patient may not be anaemic.
• May be neurological symptoms only, including
dementia.
• Levels can fall in pregnancy.
• Autoantibody tests are useful (IFA/GPC).
• Schilling test of limited use.
• Trial of B12.

28. B12 levels
• >250 ng/L – normal.
• <150 (100) – probably genuine deficiency.
• 200 -250 – borderline; repeat level.
• 150-200 – possible deficiency:
– Clinical circumstances;
– Diet (?vegan);
– Repeat/sequential levels;
– ?antibody studies.

29. Folate deficiency
• Serum folate sensitive to recent dietary
changes (eg anorexia/fasting).
• >4 genuine deficiency unlikely.
• <2 (?<3) genuine deficiency.
• 3-4 borderline; repeat.
• MCV may be helpful if >115.
• Think of malabsorption (eg Coeliac):
– Consider tTG.

30. Isolated macrocytosis
• Excess alcohol; consider GGT.
• Chronic liver disease; check albumin.
• Pregnancy.
• Myelodysplasia.
• Drugs:
– Hydroxyurea (often >120)
– Methotrexate
– Azathioprine

31. Case study 4
• 60 year old Caucasian male.
• c/o breathlessness.
• Hb= 9.0.
• MCV = 88.
• WCC and platelets normal.

32. Case study 4
• Investigations:
• Haematinics normal.
• TSH normal.
• LFTs normal.
• ESR 30.
• Creatinine 250.
• Possible cause of
anaemia?
• 2o to renal impairment
• Additional tests:
• EPO level = 15.
• Ferritin
• Retics 30 (<1%).
• Diagnosis
– Anaemia secondary to
chronic renal failure.
• Treatment
– EPO 60u/kg x1/week.
– +/- iron.

33. Normocytic anaemia
• Non-specific.
• Most non-haematinic deficiency causes.
• Can be seen with iron deficiency.
• Combined deficiencies.
• Recent bleeding.
• Anaemia of chronic disease; can be microcytic.

34. Anaemia evaluation
• Initial tests
• FBC:
• Hb (severity)
• MCV (type)
• WCC/platelets
– Bone marrow disease
– Megaloblastic anaemia
• Blood film exam
• Subsequent tests
• Reticulocyte count
– Inc. with haemolysis
– Dec. with marrow disease
or suppression
• Haematinics
– Esp. with dec/inc. MCV
• Creatinine
• LFTs
• CRP
• Protein electrophoresis
• TSH

35. Functional anaemia classification
• Acute
• Chronic  iron deficiency
Increased
loss
• RBC content (Hb; enzymes)
• RBC membrane (HS)
• RBC environment (antibodies)
Increased
destruction
• Normal marrow (deficiencies; ACD; ARF)
• Abnormal marrow (Leuk; MDS; metastases)
Decreased
production

36. End of part one!
ANY QUESTIONS?

37. Polycythaemia
• Suspect if:
– Men – Hb >18.5/HCT >52%
– Women – Hb >16.5/Hct >48%
• Persistent.
– Always repeat the FBC.

38. Polycythaemia - causes
• Primary (PRV/PV)
• Itching (bath/shower)
• Splenomegaly
• Raised WCC/platelets
• Raised uric acid/gout
• Secondary
• Heavy smoking/COPD
– Oxygen saturation
– At rest and post exercise
• Sleep apnoea/obesity
– h/o snoring
– Day time sleepiness
• Renal lesions – esp. Ca
– Abdominal USS

39. Polycythaemia – new tests
• Erythropoietin
– Low with PRV
– Raised with:
• COPD
• Renal tumours
• Not always clear cut.
• JAK2 mutations
• Janus Kinase
• Associated with cell
signaling pathways
• Mutations associated
with increased cell
proliferation
• ~95% of PRV
• ~50% PT/MF
• Normal with other causes
of polycythaemia

40. Neutrophilia
• Infection/inflammation
• Malignancy
• Myeloproliferative diseases:
– PRV/1oThrombocythaemia/Myelofibrosis
– Raised Hct or platelets; splenomegaly
• Steroid treatment
• Chronic myeloid leukaemia

41. Neutrophila – blood films
Reactive neutrophilia Chronic myeloid leukaemia

42. Eosinophilia
• Common
• Drugs
• Allergy
– Asthma; eczema;
urticaria
• Parasitic infection
• Less common
• CTDs – esp. PAN
• Some skin diseases:
– Pemphigus; DH
• Malignancy:
– Hodgkin’s disease
• Sarcoidosis
Eosinophil

43. Case study 5
• 75 year old Caucasian
male
• Routine FBC
• Hb =14
• WCC =20
• Neutros = 5
• Lymphs = 15
• Platelets = 300
• Blood film:
• Mature lymphocytes
• Smear cells

44. Case study 5
• Likely diagnosis?
• Chronic lymphatic
leukaemia.
• What physical signs
would you look for?
• Enlarged lymph nodes
or spleen.
• Additional tests
• Immunoglobulins
• Direct Antiglobulin Test
• Β2microglobulin
• Immunophenotyping
• Cytogenetics
• Bone marrow -no

45. Lymphocytosis
• Infections; esp. viral
– Age of patient
– Blood film
• Lymphoproliferative diseases:
– CLL
• Age; enlarged lymph nodes or spleen; blood film
– Lymphomas
– Hairy cell leukaemia

46. Lymphocytosis – blood films (1)
Reactive lymphocytes CLL

47. Lymphocytosis – blood films (2)
Lymphoma cells Hairy cells

48. Case study 6
• 40 year old Afro-
Caribbean male
• Routine FBC for HT.
• Hb = 14
• WCC = 2.8
• Neutros = 1.0
• Platelets = 300
• Relevant history/exam?
• Recurrent infection or
mouth ulcers – no.
• Drugs – Losartan for HT.
• Rheumatological
symptoms – no.
• Enlarged spleen – no.
• Likely diagnosis?
• Benign ethnic
neutropenia.

49. Neutropenia
• Benign-ethnic (Afro-Caribbean).
• Infections; esp. viral. ?age.
• Drugs/recent chemotherapy.
• Bone marrow disease (Hb; platelets).
• Megaloblastic anaemia (Hb/MCV).
• Immune:
– Autoimmune
– SLE
– RA (Felty’s syndrome; associated splenomegaly).

50. Lymphopenia
• SLE/RA
• HIV
• Chemotherapy
• Malignancies – eg Hodgkin’s disease
• Sarcoidosis

51. Case study 7
• 35 year old Caucasian
female
• c/o joint pains
• Hb = 12
• WBC = 4
• Neutros = 2
• Platelets = 80
• MPV = 12
• No h/o excess bleeding
• Possible diagnoses:
• Rheumatological
– SLE
• Immune – ITP
• Drugs
• HIV

52. Thrombocytopenia
• Isolated:
• Drugs
• Autoimmune – ITP
• Immune secondary to:
– SLE; LPDs.
• HIV and other
immunodeficiency
syndromes
• Ch. Liver disease with portal
HT
• Alcohol (?MCV)
• Antiphospholipid syndrome
• Other cytopenias:
• Megaloblastic anaemias
– Hb and MCV
• Bone marrow disease
– MDS
– Leukaemias
– Aplastic anaemia

53. Thrombocytopenia - investigations
• ANA
• Antiphospholipid tests:
– Anticardiolipin antibodies
– Lupus anticoagulant tests
• HIV test; esp. in at risk individuals
• LFTs
• B12 and folate

54. Thrombocytopenia in pregnancy
• Gestational
– Mild
– 2nd and 3rd trimester
• Associated with PIH syndromes
• Autoimmune (ITP)
– Often seen in 1st trimester
• HIV

55. Thrombocytosis
• Reactive:
• ?recent change
• Infection/inflammation
– ESR/CRP
• Malignancy
• Iron deficiency/bleeding
• Post splenectomy
• Primary (MPDs):
• Persistent increase
• 1oThrombocythaemia
• PRV
• Myelofibrosis
– Enlarged spleen
• CML
– Low Hb
– Raised WCC
– Splenomegaly

56. Case study 8
• 70 year Caucasian female
• c/o back pain
• FBC - normal
• U and E – normal
• LFTS - raised globulins 42
• Protein electrophoresis
showed IgG paraprotein 5
• Other Igs (A and M)
normal
• Possible diagnoses:
• Myeloma?
• “benign”?
– MGUS
• Further investigations:
• Urine for BJP - negative
• X-rays – OA changes only
• Calcium – normal
• Likely diagnosis?
• MGUS.

57. Raised globulins
• Often a non-specific reaction to infection or
inflammation.
• Protein electrophoresis usually shows a
diffuse increase in gamma globulins.
• Think of HIV in risk groups.
• Electrophoresis may show a paraprotein.

58. Protein electrophoresis
• Paraproteins:
• Myeloma (IgG or A).
• Lymphoproliferative
diseases (IgM, G or A):
– NHL
– Waldenström’s (IgM)
– CLL
• Amyloidosis.
• MGUS.
1 = Normal
2 = Diffuse increase in globulins
3 & 4 = paraproteins

59. Myeloma Vs. MGUS
• Myeloma
• New bone pain
• End organ failure
– Anaemia
– Increased creatinine
– Hypercalcaemia
• Higher concentration
paraprotein
• Immune paresis
• Bence Jones protein
• Lytic lesions on X-rays
• Typical bone marrow findings
• MGUS
• Asymptomatic
• No end organ failure
• Low concentration paraprotein
• No immune paresis
• No/minimal BJP
• No skeletal lytic lesions
• Bone marrow not always
necessary

60. Myeloma bone lesions

61. Myeloma bone marrow

62. THANKS!
ANY QUESTIONS?