CBC, interpretation, hematology, red blood cells, pediatric

1. INTERPRETATION
OF CBC
DR. N. BAJAJ

2. TERMS
 Anisopokilocytosis: variation in size and shape
 Cytometry: measurment of the cell either visual or automated
 Cluster analysis: analysis that is based upon the instrument‟s ability to cluster
different populations, together based upon size, staining, absorption or other
parameter
 Contour grating: analysis where information is plotted three dimentionally, that can
be separate subpopulation of cells
 Coulter principle (electrical impedence): sizing and counting cells by detecting and
measuring changes in electrical resistance when cell passes through small aperture.
 Dimorphic : two population of cells in single blood sample
 Forward angle light scatter: light from laser source is scatter in forward direction (0
degree) when it strike a cell or particle, larger object more forward light scatter
 Forward high angle light scatter: similar to forward angle light scatter, but angle is 5
to 15 degree variation
 Forward low angle light scatter: similar to forward angle light scatter, but angle is 2
to 3 degree variation

3. Introduction
 Haematology comprise of
 „Haima‟ = blood in Greek
 „Logos‟ = study
 Haematology is a unique super speciality in medicine
which encompasses the fields of
pathology, physiology, biochemistry, molecular
biology, obstetrics and gynecology, medicine and
paediatrics

4. CBC
 A complete blood count is a series of tests used to
evaluate the composition and concentration of the
various cellular component of the blood.
 Is a basic test
 Most informative single investigation
 Tests consists of
1. Counts of RBC, WBC, Platelets
2. Haemoglobin, haematocrit, and red cell indices
3. TLC, DLC
4. Platelet count, mean platelet volume, plateletcrit,
PDW
5. Histogram of RBC, WBC, Platelets

5. How important is CBC ?
 To know the importance of CBC we need to know…..
 What is CBC?
 Why CBC?
 What are various parameter of CBC?
 What are variation in parameter of CBC?
 What these variation can tell us?
 How these variations affect the assessesment and care of
patients?

6. Why CBC?
 CBC is an inexpensive tool and powerful
tool which provide information about
 Blood, also about
 Marrow,
 Health or disease state of other organ
of body

7. CBC USES
To diagnose
1. Anemia
2. Haemoglobinopathies
3. Bone marrow aplasia
4. Nutritional deficiencies
5. Thrombocytopenia
6. Autoimmune conditions
7. Infections and Parasitemia
8. Malignancies, response to drug, chemotherapy etc.

8. Red blood cells

9.  RBC produced in marrow and requires
 Iron, copper, magnease, cobalt
 Vitamins; especially B12, folic acid
 Regulated by erythropoietin, thyroid
hormone, androgens
 Counts depend upon
age, sex, altitude, exercise, drug, tobacco
use etc.
 Life span - 120 days

10.  Clinical importance of assessment of RBC is to: measures oxygen
carrying capacity of blood
Normal values
 Newborn 4.1-6.1 million/mm3
 Children 3.6-5.5 million/mm3
 Adult (M) 4.6-6.0 million/mm3
 Adult (F) 4.2-5.0 million/mm3

11. Decreased RBCs
Blood loss Impaired production Increased destruction
• Trauma
• Surgery
• GI bleed
• Gynecological
disturbance
• Pure red cell aplasia
• Pernicious anemia
• Megaloblastic anemia
• Iron deficiency anemia
• Thalassemia
• Anemia of prematurity
• Anemia of chronic
disorder
Intra-corpuscular
• Hereditary spherocytosis
• Sickle cell anemia
• Abetalipoprotienimia
• G6PD
• Pyruvate kinase deficiency
• PNH
Extra-corpuscular
• Autoimmune
• Haemolytic disease of newborn
• Mismatch transfusion
• Microangiopathic haemolytic
anemia TTP, HUS
• DIC
• infections

12. • Increased RBCs
• Polycethemia vera
• High altitude
• chronic obstructive pulmonary disease(COPD,
emphysema, chronic bronchitis),
• pulmonary hypertension,
• Hypoventilation syndrome,
• congestive heart failure
• obstructive sleep apnea,
• poor blood flow to the kidneys, and

13. Haemoglobin
 Oxygen carrying component of blood
 Synthesize in polychromatic normoblast stage of red cell development
 Values
 Newborn 15.5-24.5 g/L
 Adult male 13.5-16.5 g/L
 Adult female 12.0-15.5 g/L

14. Hb estimation
Cynemethamoglobin Method: Recommended
 20 microL blood + diluent (potassium cynide
and potassium fericynide)
 Mixed and read in photo colorimeter
 Photo colorimeter is used to determine the
concentration
 Hb% =(test sample absorbance/ standard sample
absorbance)x concentration of standard x dilution
factor
 Advantage – haemoglobin, methamoglobin and
carboxyhaemoglobin are used in measurment.
 Disadvantage – sulphamethamoglobin cannot be
included in measurment, takes more time for
estimation

15. Hb estimation: other method
 Sahli acid haematin
 Alkaline haematin method
 Sulphahaemoglobin method
 Oxyhaemoglobin method

16. Reticulocyte
 Normal value 0.5% - 1.5%.
Hence 0.5% - 1.5% RBCs are replaced per day
 Uses
 To evaluate anemia
 Response to treatment of anemia
 Note
 If the disease causing the anemia is inside the
marrow, the reticulocyte count is decreased
 If the disease causing the anemia is outside the
marrow, the reticulocyte count is increased

17. Methods
 Manual reticulocyte count using supravital stain
 Automated reticulocyte count by flouroscent
method - gives immature reticulocyte fraction
(IRF) and removes errors like Howell-Jolly
bodies, pappenheimer bodies
 Reticulocyte production index or corrected
reticulocyte count: an index corrected according
to level of anemia
 Reticulocyte index = reticulocyte count x
patient‟s haematocrit/ normal
haematocrit

18.  Reticulocyte proliferation index: Index is used to
determine if a person's bone marrow is properly
responding to the body's need for red blood cells
 Shift correction factor: normal reticulocyte count
survive 3.5 days in marrow and 1 day in peripheral
circulation at normal PCV. In case of variation in PCV
the survival time is increased which is termed as shift
correction factor
 Reticulocyte proliferation index = reticulocyte
index/ shift correction factorShift correlation factor
PCV% Maturation days = shift correction factor
45 1
35 1.5
25 2
15 2.5

19. Increased reticulocyte count
 Haemolytic anemia
 Recent haemorrhage
 Thalassemia
 Pregnancy
 Response to treatment
 Hypoxia
 Leukamia
Decrease reticulocyte count
 Aplastic anemia
 Megaloblastic anemia
 Anemia of chronic disease
 Cirrhosis
 Radiation
 Decrease ACTH and pitutary
hormones

20. Reticulocyte haemoglobin
measurement (RET-He)
 Reticulocyte Hemoglobin (Ret-He) is a direct assessment of the incorporation
of iron into erythrocyte hemoglobin.
 It is a direct estimate of the recent functional availability of iron (2–3 days).
 Traditional chemistry tests used for iron assessment (serum iron, Tsat,
ferritin) are indirect measurements.
 As a direct measurement, Ret-He may identify iron deficiency earlier than
traditional parameters.
 It is an established parameter used in KDOQI (Kidney Disease Outcome
Quality Initiative) guidelines for assessing iron status

21. Haematocrit
 Ratio of the volume of erythrocytes to that of the whole blood in percentage
 Most precise method for determining the degree of anemia or
polycythemia i.e. increase or decrease RBC concentration
Normal values
 Newborn 42-68%
 Upto 1 year age 29-41%
 Adult Male 39-47%
 Adult female 36-44%
 Rule of 3:– RBC x 3 = Hb and Hb x 3 = Hct

22. High
 Polycythemia vera
 Dehydration
 Low oxygen in blood
 Congenital heart disease
 Cor pulmonale
 Smoking
 Haemoconcentration (Dengue)
Low
 Anemia
 Blood loss
 Haemolysis
 Bone marrow aplasia
 Leukamia
 Malnutrition
An elevated haematocrit may be due to spleen hyper
function, and reduced haematocrit may indicate low thymus
function

23. Mean corpuscular volume
 Measures average volume of RBC
 MCV = haematocrit/ red cell count x100
 Normal values
 Newborn 103-106fL
 Child upto 1 year 78 fL
 Adult 79-98fL
 Classified accordingly as
 Microcyte – MCV <79
 Macrocytic – MCV >98
 Presence of microcytic and macrocytic cells in same sample may result
in normal MCV
 MCV <72 without heterogeneity, is a sensitive and specific predictor
of thalassemia trait

24. Microcytic MCV
 Hypochromic
 Iron deficiency
 Thalessemia
 Lead poisoning
 Porphyria
 Normochromic
 Anemia of chronic
disease
 haemoglobinopath
ies
 Macrocytic MCV
 Megaloblastic
anemia
 Pernicious anemia
 Sprue
 Di Gulielmo disease
 MDS
 Post spleenectomy
 Alcoholism
 Liver disease
 Drugs
(anticonvulscents,
anticancer etc)
• Normocytic MCV
• Acute haemorrhage
• Diamorphic anemia
• Haemoglobinopathi
es
• Endocrinopathies

25. Interference in MCV
 Cold and warm antibodies
 Marked hyperglycemia
 Marked leukocytosis
 Marked reticulocytosis
 Methanol poisoning

26. Mean corpuscular
haemoglobin
MCH = haemoglobin/ red cell count x 100
Normal range
 Newborn 36-38%
 Upto 1 year age 23-27%
 Adult 26.7-31.9%

27.  MCH decreased in
 Microcytic and normocytic anemias
 MCH increased in
 Macrocytic anemias
 Infants and newborns
 Interference in MCH
 Lipemia
 Marked leukocytosis
 Cold agglutinin
 Monoclonal protein in blood

28. Mean corpuscular
haemoglobin concentration
MCHC = haemoglobin/ haematocrit x 10
Normal range
 Newborn 34-36%
 Upto 1 year age 31-33%
 Adult 32-36%

29.  MCHC decreased in
 Hypocromic microcytic anemia
 MCHC increased in
 Heridietery spherocytosis
 Infant and newborns
 Autoagglutinations
 Interference in MCHC
 Marked leukocytosis
 Haemolysis
 Cold aggutinins
 Rouleaux

30. Red cell distribution width
(RDW)
 Red cell distribution is a quantative measure or numerical
expression of anisocytosis. It is a coefficient of variation of the
distribution of individual RBC volume
 In microcytes, RDW increased in iron deficiency anemia but in
thalessemia it is not raised

31.  RDW-CV:It is the ratio of standard deviation to the
mean corpuscular volume
RDW-CV = standard deviatiom of RBC volume/ mean MCV x
100
value 11.5%-14.5%
 RDW-SD: It is the actual measurnment of the width of
the RBCdistribution curve
 Values 35-45 fL

32. RBCs on peripheral smear

33. Preparation
 The wedge slide (push slide) technique was developed
by Maxwell Wintrobe as is a standard method
 The “zone of morphology” (area of optimal thickness for
light microscopy examination) should be at least 2cm in
length. The smear should occupy the central area of the
slide and be margin free at the edges.

34. RBCs in peripheral smear
 Microcytic hypochromic
 Size smaller than the nucleus of small
lymphocyte
 < 7 micron
 Markedly increase central pallor >1/3
of the diameter of RBC
 Causes
 Iron deficiency anemia
 Thalassemia
 Sideroblastic anemia
 Anemia of chronic disease
 Haemoglobinopathies

35. Macrocytic cells
 Size > 8.3 micron kin diameter
 Causes
 Vitamin B 12 and folic acid deficiency
 Alcoholism
 Liver disease
 Myleodysplastic syndrome
 Hypothyroidism
 Drug that impair DNA synthesis

36.  Oval macrocytes
 Vitamin B 12 and folic acid deficiency
 Pernicious anemia
 Myleodysplastic syndrome
 Hypothyroidism
 Drug that impair DNA synthesis

37.  Round hypochromic macrocytes
 Alcoholism
 Hypothyroidism
 Liver disease
 Post splenectomy
 Blue tinged macrocytes
 Neonate
 Response to anemic stress

38. Target or bell cell
 They have a characteristic ringed
appearance. This is because of the
“increase surface area to volume ratio”
i.e. increase in red cell membrane which
get pooled at the centre of cells
 Causes
 Thalessemia
 Haemoglobinopathies Hb AC or CC,
HbSS,SC
 Liver disease
 Post spleenectomy
 Severe iron deficiency anemia
 abetalipoprotenimia

39. Schistocytes
 „Schisto‟ = split or cleft
 Physical assault to erythrocytes with in the
blood stream creates these cells
 which include
 Helmet cells
 Triangles
 Crescents
 Microspherocytes
 Horns
 Purse

40. Causes
 DIC
 Severe haemolytic anemia
 Microangiopathic haemolytic anemia
 HUS &TTP
 Prosthetic cardiac valves
 Connective tissue disorders
 Burns
 Acute tubular necrosis, glomerulonephritis
 Malignant hypertension

41. Tear drop cells(Dacrocytes)
 Pear shape cells, usually microcytic and hypochromic
 Seen in
 Newborn
 Thalassemia major
 Myleoproliferative disorder
 Leukoerythroblastic reaction

42. Spherocytes
 Ball shaped red cells, decreased surface: volume
ratio, hyperdense (> MCHC)
 Seen in
 Hereditary spherocytes
 ABO incompatibility
 Autoimmune haemolytic anemia
 Microangiopathic haemolytic anemia
 SS disease
 Hyperspleenism
 Burns
 Posttransfusion

43. Elliptocytes
 Elliptical and normochromic cells, seen normally in
less than 1% of RBCs
 Causes
 Hereditary elliptocytosis
 Iron deficiency anemia (increased with severity)
 SS disease and SA trait
 Thalassemia Major
 Leukoerythroblastic reaction
 Malaria
 Megaloblastic anemia

44. Burr cells (Echinocytes)
 10-30 spicules equal in size and evenly distributed over
RBC surface; caused by alteration in extracellular
environment
 Seen in
 Liver disease
 Renal failure
 Dehydration
 Pyruvate kinase deficiency
 Storage artefacts

45. Spur cell - Acanthocyte
 Acantho = thorn
 Cells with 5-10 specules of varying length, irregular in
shape, thickness, with wide bases and appear smaller than normal
cell because they assume spheroid shape
 Result from changes in membrane lipid content
 Seen in
 Spur cell anemia
 Alcoholism
 Hypothyroidism
 Abetalipoprotinemia
 Vitamin E deficiency
 Malsbsorption
 Postsplenectomy

46. Bite cell (Degmacyte)
 Appear as a cookie with a bite taken
out
 Seen in G6PD
 When spleen removes the Heinz bodies
from RBCs

47. Stomatocyte
 When examined on dry smear, it has a central slit or stoma
 Seen in
 Few may be seen normally
 Various cardiovascular
and pulmonary disorders
 Hereditary
 Alcoholism
 Liver disease
 Malignancies

48. Howell – Jolly bodies
 Small well defined, rounded, densely stained
inclusions, 1 micron in diameter, ecentric, that
represent DNA fragments
 Associated with rapid or abnormal RBC formation
 Seen in
 Post spleenectomy
 Newborns
 Megaloblastic anemia
 Dyserythopoietic anemias
 Hereditary spherocyosis

49. Heinz Bodies
 Inclusion of denatured haemoglobin caused by oxidation
of globin portion of haemoglobin
 Removal of Heinz body leads to formation of „bite cells‟
 Causes
 Drugs
 Certain foods like fava beans
and onion

50. Sideroblastic granules/
pappenheimer bodies
 Irregular dark blue iron containing granules occuring in
small clusters, predominantly in periphery
 Seen in
 Sideroblastic anemia
 Spleenectomy
 Haemolytic anemia
 Myelodysplastic syndromes
 Lead poisoining
 Its presence can rule out iron deficiency anemia

51. Sickle cell
 Crescent shape cells develop in
 people homozygous for haemoglobin S
 Heterozygous HbS and either
thallasemia or another Hb like Hb C

52. Nucleated red cells
 Cells have dense dark nucleus in the center
of the cell
 Results from marked stimulation of the
bone marrow
 Seen in
 New born (first 3-4 days)
 Acute bleeding severe haemolytic anemia
 Megaloblastic anemia
 Congenital infections (syphilis, CMV, rubella)
 Postspleenectomy
 Leukoerythroblastic reaction
 Fungal and mycobacterial infections
 Dyselectropoeitic anemia

53. Basophilic stippling
 Numerous small, purplish inclusions, which results from
RNA and mitochrondrial remenants
 Seen in
 Lead toxicity
 Thalessemia
 Haemoglobinopathies
 Macrocytic anemia

54. Cabot ring
 These are delicate thread like inclusions, remenants of
the nuclear membranes, in the RBC
 They can take any shape like purplish ring, figure of
eight, incomplete ring
 Seen in
 Pernicious anemia
 Lead poisoning
 Alcoholic jaundice
 Severe anemia
 Leukamia

55. Roulex formation
 A stack like arrengment of red blood cells where the
biconcave surdface of RBCs are next to each other.
 Seen in
 Increase in cathodal protien,
such as immunoglobins
and fibronegen
 Multiple myleoma
 Macroglobulimias
 Acute and cronic infections
 Connective tissue disease
 Diabetes mellitus
 Malignancies

56.  Grading of inclusions
 Rare 0-1/hpf
 Few 1-2/hpf
 Mod 2-4/hpf
 Many >5/hpf
 Qualitative grading of abnormal RBC morphology
Grade degree of abnormalities
1-5 cells /10fields slight
6-15cells /10fields moderate
>15cells /10fields marked