interpretation of cbc
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INTERPRETATION OF CBC
DR. N. BAJAJ
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
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
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
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?
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
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.
Red blood cells
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
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
Decreased RBCsBlood loss Impaired production Increased destruction
• Trauma • Surgery• GI bleed• Gynecologic
al 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• PNHExtra-corpuscular• Autoimmune• Haemolytic disease of
newborn• Mismatch transfusion• Microangiopathic haemolytic
anemia TTP, HUS• DIC• infections
• 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
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
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
Hb estimation: other method
Sahli acid haematin
Alkaline haematin method
Sulphahaemoglobin method
Oxyhaemoglobin method
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
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
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 factor
Shift correlation factor
PCV% Maturation days = shift correction factor
45 1
35 1.5
25 2
15 2.5
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
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
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
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
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
Microcytic MCV Hypochromic
Iron deficiency
Thalessemia
Lead poisoning
Porphyria
Normochromic Anemia of
chronic disease
haemoglobinopathies
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
• Haemoglobinopathies
• Endocrinopathies
Interference in MCV
Cold and warm antibodies
Marked hyperglycemia
Marked leukocytosis
Marked reticulocytosis
Methanol poisoning
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%
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
Mean corpuscular haemoglobin concentration
MCHC = haemoglobin/ haematocrit x 10
Normal range
Newborn 34-36%
Upto 1 year age 31-33%
Adult 32-36%
MCHC decreased in
Hypocromic microcytic anemia
MCHC increased in
Heridietery spherocytosis
Infant and newborns
Autoagglutinations
Interference in MCHC
Marked leukocytosis
Haemolysis
Cold aggutinins
Rouleaux
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
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
RBCs on peripheral smear
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.
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
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
Oval macrocytes
Vitamin B 12 and folic acid deficiency
Pernicious anemia
Myleodysplastic syndrome
Hypothyroidism
Drug that impair DNA synthesis
Round hypochromic macrocytes
Alcoholism
Hypothyroidism
Liver disease
Post splenectomy
Blue tinged macrocytes
Neonate
Response to anemic stress
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
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
Causes
DIC
Severe haemolytic anemia
Microangiopathic haemolytic anemia
HUS &TTP
Prosthetic cardiac valves
Connective tissue disorders
Burns
Acute tubular necrosis, glomerulonephritis
Malignant hypertension
Tear drop cells(Dacrocytes)
Pear shape cells, usually microcytic and hypochromic
Seen in
Newborn
Thalassemia major
Myleoproliferative disorder
Leukoerythroblastic reaction
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
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
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
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
Bite cell (Degmacyte)
Appear as a cookie with a bite taken out
Seen in G6PD
When spleen removes the Heinz bodies from RBCs
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
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
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
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
Sickle cell
Crescent shape cells develop in
people homozygous for haemoglobin S
Heterozygous HbS and either thallasemia or another Hb like Hb C
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
Basophilic stippling
Numerous small, purplish inclusions, which results from RNA and mitochrondrial remenants
Seen in
Lead toxicity
Thalessemia
Haemoglobinopathies
Macrocytic anemia
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
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
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
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