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Peripheral blood smear examination
Dr Shanaz KhodaijiConsultant Hematopathologist
P.D. Hinduja National Hospital & Medical
Research Centre
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Hemogram:
measured and
calculated
parameters
Histograms:
size distribution
of WBC, RBCand Plt
Cytogram:
WBC differential
CBC on automated analyzers
Flagging for abnormalitiesnecessitates a manual PBS
review
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A well made peripheral smear is thick at one end and progressively thinnerat the opposite end. The "zone of morphology" (area of optimal thicknessfor light microscopic examination) should be at least 2 cm in length. Thesmear should occupy the central area of the slide and be margin-free at
the edges
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PBS examination requires a systematic approach in
order to gather all possible information. In addition, allspecimens must be evaluated in the same manner, toassure that consistent information is obtained.
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The examination starts with
Macroscopic view to evaluate the quality of the smear
The microscopic analysis begins on lower power (10x),
primarily to assess cellular distribution, staining quality,
and to select an area where the RBCs are barely touching
each other. This area is used to assess the cellular
elements on higher magnification.On hi-dry (40x), the slide is principally scanned to obtain a
WBC estimate. All of the detailed analysis of the cellular
elements is performed using high power or oil immersion.
PBS examination - preliminary
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(a) Ten microscopic fields are examined in a vertical directionfrom bottom to top or top to bottom (b) The slide ishorizontally moved to the next field (c) Ten microscopic fieldsare counted vertically. (d) The procedure is repeated until100 leukocytes have been counted
Scanning technique for WBC differentialcount and morphologic evaluation
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This final microscopic examination is performed at 50x
or 100x oil immersion and includes:
A WBC differential
The identification of abnormal leukocytes
Assessment of RBC morphology
The number and morphology of the platelets
The identification of intra- and extra-cellular elements Assessment of any organisms present
PBS examination - final
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Microscopic examination of the peripheral blood is used to
supplement information provided by CBC analyzers.
Hematology analyzers provide accurate quantitative
information about blood cells and can identify abn cells
In addition to providing cell counts and graphical displays
these instruments also provide a warning flags
The instrument operator reviews the information from each
specimen and decides if smear preparation and light
microscopy are necessary.
If not, the information is released to the clinician.
Hematology analyzer and PBS
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A fairly accurate estimate of the WBC count (cells/mL)
can be obtained by counting the total number of
leukocytes in ten 50X microscopic fields, dividing the
total by 10, and multiplying by 3000. These estimates
should approximate that obtained by the cell analyzer.
If the estimate does not match the automated cell
count, obtain the original blood specimen, confirmpatient identity, repeat the automated analysis, and
prepare a new smear.
WBC estimation on peripheral smear
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Morphologic Evaluation of Red Blood Cells
Round to very slightly ovoid
cells with a mean diameter of
approximately 7 A central pale area - central
pallor approximately 1/3 thediameter of the cell It is approximately the same
size as the nucleus of a
mature lymphocyte. Any deviation in size, volume,
or shape represents an
abnormal red blood cell.
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Microcytic hypochromic red cells
Decreased size and Hbcontent (MCH) and conc
(MCHC). Expanded
central zone of pallor
Iron deficiency, thal trait? Anemia of chronic
disease, sideroblastic
an
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Iron deficiency anemia
Tanja Tornow
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CBC + reticulocyte count in ACD
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Elliptocytes or ovalocytes
Ovalocytes are due to abnormal membranecytoskeleton found in hereditary elliptocytoisis
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Megaloblastic anemia (PS)
Macrocyte
"Thin"
macrocyte
Large RBCs (> 8.5
mm, MCV > 95 fL).Normal MCH
Increased diameter,
normal MCV. Usuallyhypochromic
Accelerated
erythrocytosis.Macrocytic anemia
(B12/folate def) (oval
macrocytes)
Liver disease,postsplenectomy
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Tear drop shaped red cells or dacryocytes areseen when there is extramedullary erythropoiesisor with marrow disorders or marrow infiltration,such as myelofibrosis or metastatic carcinoma.
Tear drop cells / dacrocytes
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Polychromasia
Blue-gray coloration
of RBCS. Due RNA
remnants
Increased - Increased
erythropoietic activity.
Decreased -
Hypoproliferative states.
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Sickle cell anemia
Irregular, curved
cells with pointed
ends
Hb S hemoglobinopathies (sickle cell
anemia, hb SC disease, hb S-beta-
thalassemia, hb SD disease, hb
Memphis /S disease), other
hemoglobinopathies (especially Hb I, Hb
CHarlem, HbCCapetown).
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Spherocytosis
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Acanthocytes or spur cells, are
spherical cells with blunt-tipped or
club-shaped spicules of different
lengths projecting from their surface
at irregular intervals. (Echinocytes,
or crenated red cells, in contrast,
have shorter, sharp to blunt
spicules of uniform length which are
more evenly spaced around their
periphery).
Acanthocytes
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Spheroid RBCswith few large
spiny projections.
5-10 spicules,
irregular spacing
and thickness
(must be
differentiated fromechinocytes).
Abetalipoproteinemia,postsplenectomy, alcoholic
cirrhosis and hemolytic anemia,
microangiopathic hemolytic
anemia, autoimmune hemolytic
anemia, sideroblastic anemia,
thalassemia, severe burns, renal
disease, pyruvate kinasedeficiency, McLeod phenotype,
infantile pyknocytosis,
Acanthocytes or spur cells
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Echinocytes
"Sea urchin
cells,
crenated cells,burr cells"
RBC with
many tiny
spicules (10-
30) evenlydistributed
over cell
Post-splenectomy, uremia,
hepatitis of the newborn,
malabsorption states, after
administration of heparin,pyruvate kinase def
phosphoglycerate kinase
deficiency, uremia, HUS.
Crenated / Burr cells / Echinocytes
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Mechanical damage to
RBCs from fibrin deposits
DIC
MAHA
TP prosthetic heart valves
severe valvular stenosis
malignant hypertension march hemoglobinuria
myelofibrosis hypersplenism
Schistocyte fragmented RBC
normal newborns
bleeding peptic ulcer
Aplastic Anemia
pyruvate kinase def
VasculitisGlomerulonephritis
renal graft rejection
severe burnsiron deficiency, thalassemia
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Some specific hemolyic anemias
Mechanical HAs: RBC are injured by excessphysical trauma as they circulate in the bloodvessels
Cardiac hemolytic anemias artificial heart
valveMAHA - due to thrombosed vessels or fibrin
strands
as in DIC, TTP, malignancy
Hallmark: Presence of schistocytes in the PB
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Uniconcave RBC,
slitlike area of central
pallor
Hereditary or acquired hemolysis.
Hereditary stomatocytosis, alcoholic
cirrhosis, acute alcoholism,
obstructive liver disease,
malignancy, severe infection, treated
acute leukemia, artifact.
Stomatocyte fish mouth cell
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HA due to red cell enzymedefects bite or blister cells
G6PD deficiency Sex linked transmission Presents as
hemolysis after drug intake, infections
Common drugs antimalarials, acetanilide dapsone furantoinLab diagnosis screening, quantitative tests
Pyruvate kinase deficiency uncommon
Lab diagnosis fluorescent spot test
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Irregular RBC
agglutination/
clumping
Anti-RBC antibody, paraprotein. Cold
agglutinin disease, autoimmune
hemolytic anemia,
macroglobulinemia,
hypergammaglobinemia
RBC autoagglutination
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Small (1 mm),
round, dense,
basophilic bodiesin RBCs.
Splenectomized patients,
megaloblastic anema, severe
hemolytic processes,hyposplenism, myelophthistic
anemia.
Howell Jolly bodies
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WBC Morphology
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Large, coarse, dark purple, azurophilic granules that occur in
the cytoplasm of most granulocytes. These arecharacteristically found in the Alder-Reilly anomaly and inpatients with mucopolysaccharidoses
Alder-Reilly anomaly
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Chdiak-Higashi granules are very large red or bluegranules that appear in the cytoplasm of granulocytes,lymphocytes, or monocytes in patients with the Chdiak-Steinbrinck-Higashi syndrome. It is a rare autosomalrecessive disorder
Chdiak-Higashi
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Variably sized (0.1 to 2.0 um) and shaped, blue or grayish-blue cytoplasmic inclusions usually found near the periphery
of the cell. Dohle bodies are lamellar aggregates of roughendoplasmic reticulum, which appear in the neutrophils,bands, and metamyelocytes of patients with infection,burns, uncomplicated pregnancy, toxic states, or duringtreatment with hematologic growth factors - G-CSF.
Dhle bodies
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May-Hegglin anomaly
Neutrophils contain small basophilic cytoplasmic granuleswhich represent aggregated ribosomes. Leukopenia andlarge platelets are also found. An autosomal dominant trait,the May-Hegglin anomaly is associated with a mild bleedingtendency, but not by an increased susceptibility to infection
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Neutrophilic toxic granulation
Small dark blue to purple granules resembling primary
granules in the cytoplasm of metamyelocytes, bands, andsegmented neutrophils during inflammatory states, burns,
and trauma, and upon exposure to hematopoietic growth
factors. It is usually accompanied by a shift to the left and
vacuolations in the cytoplasm (toxic vacuolations) andDohle bodies.
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Macroplatelets
Platelet morphology
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Platelet satellitism
M t i ith i t l t l t (MDS
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Macrocytosis with giant platelets (MDS,5q- syndrome)
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Disadvantages of the Peripheral Blood Smear
Provides information that cannot be obtained from automated
cell counting. However, some limitations are:
Experience is required to make technically adequate smears.
There is a non-uniform distribution of white blood cells over
the smear, with larger leukocytes concentrated near the edges
and lymphocytes scattered throughout.
There is a non-uniform distribution of RBCs over the smear,
with small crowded red blood cells at the thick edge and large
flat red blood cells without central pallor at the feathered edge
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Thank You